Benzimidazoles for inhibiting protein tyrosine kinase mediated cellular proliferation

ABSTRACT

Benzimidazoles of Formula I below are inhibitors of protein tyrosine kinases, and are useful in treating cellular proliferation. ##STR1## The compounds are especially useful in treating cancer, atherosclerosis, restenosis, and psoriasis.

CROSS REFERENCES TO RELATED APPLICATIONS

This application claims priority of U.S. Ser. No. 60/056,609, filed Aug.20, 1997.

FIELD OF THE INVENTION

This invention relates to the inhibition of protein tyrosine kinase(PTK) mediated cellular proliferation. More specifically, this inventionrelates to benzimidazoles and their use in inhibiting cellularproliferation and protein tyrosine kinase enzymatic activity.

BACKGROUND OF THE INVENTION

Many disease states are characterized by the uncontrolled proliferationand differentiation of cells. These disease states encompass a varietyof cell types and maladies such as cancer, atherosclerosis, restenosis,and psoriasis. Growth factor stimulation, autophosphorylation, and thephosphorylation of intracellular protein substrates are importantbiological events in the pathomechanisms of proliferative diseases.

In normal cells, the phosphorylation of tyrosine residues on proteinsubstrates serves a critical function in intracellular growth signalingpathways initiated by stimulated extracellular growth factor receptors.For example, the association of growth factors such as Platelet DerivedGrowth Factor (PDGF), Fibroblast Growth Factor (FGF), and EpidermalGrowth Factor (EGF) with their respective extracellular receptors,PDGFr, FGFr, and EGFr, activates intracellular tyrosine kinase enzymedomains of these receptors, thereby catalyzing the phosphorylation ofeither intracellular substrates or the receptors themselves. Thephosphorylation of growth factor receptors in response to ligand bindingis known as autophosphorylation.

For example, the EGF receptor has as its two most important ligands EGFand Transforming Growth Factor α, (TGFα). The receptors appear to haveonly minor functions in normal adult humans, but are implicated in thedisease processes of a large portion of all cancers, especially colonand breast cancer. The closely related Erb-B2 and Erb-B3 receptors havea family of Heregulins as their major ligands, and receptoroverexpression and mutation have been unequivocally demonstrated as themajor risk factor in poor prognosis breast cancer.

The proliferation and directed migration of vascular smooth muscle cells(VSMC) are important components in such processes as vascularremodeling, restenosis and atherosclerosis. Platelet-derived growthfactor has been identified as one of the most potent endogenous VSMCmitogens and chemoattractants. Elevated vascular mRNA expression ofPDGF-A and -B chains and PDGF receptors has been observed inballoon-injured rat carotid arteries (J Cell. Biol., 1990; 111:2149-2158). In this injury model, infusion of PDGF also greatlyincreases intimal thickening and migration of VSMC (J. Clin. Invest.,1992; 89: 507-511). Furthermore, PDGF-neutralizing antibodiessignificantly reduce intimal thickening following balloon injury(Science, 1991; 253: 1129-1132). Tyrphostin receptor tyrosine kinaseinhibitors which block the PDGF signal transduction pathway have beenshown to inhibit PDGF stimulated receptor tyrosine kinasephosphorylation in vivo in the rat cuff injury model (Drug Develop.Res., 1993; 29: 158-166).

Both acidic fibroblast growth factor (aFGF) and basic fibroblast growthfactor (bFGF) have many biological activities, including the ability topromote cellular proliferation and differentiation. Direct evidence insupport of FGF involvement in VSMC has been reported by Lindner andReidy (Proc. Natl. Acad. Sci USA, 1991; 88: 3739-3743), who demonstratedthat the systemic injection of a neutralizing antibody against bFGFprior to balloon angioplasty of rat carotid arteries inhibitedinjury-induced medial SMC proliferation by greater than 80% whenmeasured 2 days after injury. It is likely that bFGF released fromdamaged cells is acting in a paracrine manner to induce VSMC growth.Recently, Lindner and Reidy (Cir. Res., 1993; 73: 589-595) demonstratedan increased expression of both MRNA for bFGF and FGFR-1 in replicatingVSMCs and endothelium in en face preparations of balloon-injured ratcarotid arteries. The data provides evidence that in injured arteriesthe ligand/receptor system of bFGF and FGFR-1 may be involved in thecontinued proliferative response of VSMCs leading to neointimaformation.

Buchdunger, et al., Proc. Natl. Acad. Sci., Vol. 92, March 1995,2558-2562, reported the inhibition of the PDGF signal transductionpathway both in vitro and in vivo by a PDGF receptor tyrosine proteinkinase inhibitor. The compound showed antitumor activity in tumor modelsusing astrocytoma cell lines.

Thus, EGF, PDGF, FGF, and other growth factors play pivotal roles in thepathomechanisms of cellular proliferative diseases such as cancer,atherosclerosis, and restenosis. Upon association with their respectivereceptors, these growth factors stimulate tyrosine kinase activity asone of the initial biochemical events leading to DNA synthesis and celldivision. It thereby follows that compounds which inhibit proteintyrosine kinases associated with intracellular growth factor signaltransduction pathways are useful agents for the treatment of cellularproliferative diseases. We have now discovered that certainbenzimidazoles inhibit protein tyrosine kinases, and are useful intreating and preventing atherosclerosis, restenosis, and cancer.

SUMMARY OF THE INVENTION

The present invention provides a method of treating cancer, the methodcomprising administering to a patient having cancer a therapeuticallyeffective amount of a compound of Formula I ##STR2## wherein Ar is aryl,substituted aryl, heteroaryl, or substituted heteroaryl;

R¹, R², R³, and R⁴ are each independently hydrogen, C₁ -C₆ alkyl, --OC₁-C₆ alkyl, --OH, halogen, --CO₂ R^(a), --CONR^(a) R^(b), --NO₂, --NR^(a)R^(b), --COC₁ -C₆ alkyl, --CHO, --CN, --SO₂ C₁ -C₆ alkyl, ##STR3##--OCH₂ CH(OH)CH₂ OH, --O(CH₂)_(n) NR^(a) R^(b), --O(CH₂)_(n)N-morpholino, --SH, --SC₁ -C₆ alkyl, or --S(CH₂)_(n) NR^(a) R^(a), or R²and R³ together can form a cycloalkyl ring that may contain one or moreheteroatom;

R^(a) and R^(b) are each independently hydrogen or C₁ -C₆ alkyl, n is 0to 5, and the pharmaceutically acceptable salts, esters, amides, andprodrugs thereof.

The present invention also provides a method of treating or preventingrestenosis, the method comprising administering to a patient havingrestenosis or at risk of having restenosis a therapeutically effectiveamount of a compound of Formula I ##STR4## wherein Ar is aryl,substituted aryl, heteroaryl, or substituted heteroaryl;

R¹, R², R³, and R⁴ are each independently hydrogen, C₁ -C₆ alkyl, --OC₁-C₆ alkyl, --OH, halogen, --CO₂ R^(a), --CONR^(a) R^(b), --NO₂, --NR^(a)R^(b), --COC₁ -C₆ alkyl, --CHO, --CN, --SO₂ C₁ -C₆ alkyl, ##STR5##--OCH₂ CH(OH)CH₂ OH, --O(CH₂)_(n) NR^(a) R^(b), --O(CH₂)_(n)N-morpholino, --SH, --SC₁ -C₆ alkyl, or --S(CH₂)_(n) NR^(a) R^(b), or R²and R³ together can form a cycloalkyl ring that may contain one or moreheteroatom;

R^(a) and R^(b) are each independently hydrogen or C₁ -C₆ alkyl, n is 0to 5, and the pharmaceutically acceptable salts, esters, amides, andprodrugs thereof.

In another embodiment, the present invention provides a method oftreating atherosclerosis, the method comprising administering to apatient having atherosclerosis a therapeutically effective amount of acompound of Formula I ##STR6## wherein Ar is aryl, substituted aryl,heteroaryl, or substituted heteroaryl;

R¹, R², R³, and R⁴ are each independently hydrogen, C₁ -C₆ alkyl, -OC₁-C₆ alkyl, --OH, halogen, --CO₂ R^(a), --CONR^(a) R^(b), --NO₂, --NR^(a)R^(b), --COC₁ -C₆ alkyl, --CHO, --CN, --SO₂ C₁ -C₆ alkyl, ##STR7##--OCH₂ CH(OH)CH₂ OH, --O(CH₂)_(n) NR^(a) R^(b), or --O(CH₂)_(n)N-morpholino, --SH, --SC₁ -C₆ alkyl, or --S(CH₂)_(n) NR^(a) R^(a), or R²and R³ together can form a cycloalkyl ring that may contain one or moreheteroatom;

R^(a) and R^(b) are each independently hydrogen or C₁ -C₆ alkyl, n is 0to 5, and the pharmaceutically acceptable salts, esters, amides, andprodrugs thereof.

Also provided is a method of treating psoriasis, the method comprisingadministering to a patient having psoriasis a therapeutically effectiveamount of a compound of Formula I ##STR8## wherein Ar is aryl,substituted aryl, heteroaryl, or substituted heteroaryl;

R¹, R², R³, and R⁴ are each independently hydrogen, C₁ -C₆ alkyl, --OC₁-C₆ alkyl, --OH, halogen, --CO₂ R^(a), --CONR^(a) R^(b), --NO₂, --NR^(a)R^(b), --COC₁ -C₆ alkyl, --CHO, --CN, --SO₂ C₁ -C₆ alkyl, ##STR9##--OCH₂ CH(OH)CH₂ OH, --O(CH₂)_(n) NR^(a) R^(b), --O(CH₂)_(n)N-morpholino, --SH, --SC₁ -C₆ alkyl, or --S(CH₂)_(n) NR^(a) R^(b), or R²and R³ together can form a cycloalkyl ring that may contain one or moreheteroatom;

R^(a) and R^(b) are each independently hydrogen or C₁ -C₆ alkyl, n is 0to 5, and the pharmaceutically acceptable salts, esters, amides, andprodrugs thereof.

In addition, the present invention provides a method of inhibitingprotein tyrosine kinases, the method comprising administering to apatient in need of protein tyrosine kinases inhibition a proteintyrosine kinase inhibiting amount of a compound of Formula I ##STR10##wherein Ar is aryl, substituted aryl, heteroaryl, or substitutedheteroaryl;

R¹, R², R³, and R⁴ are each independently hydrogen, C₁ -C₆ alkyl, --OC₁-C₆ alkyl, --OH, halogen, --CO₂ R^(a), --CONR^(a) R^(b), --NO₂, --NR^(a)R^(b), --COC₁ -C₆ alkyl, --CHO, --CN, --SO₂ C₁ -C₆ alkyl, ##STR11##--OCH₂ CH(OH)CH₂ OH, --O(CH₂)_(n) NR^(a) R^(b), --O(CH₂)_(n)N-morpholino, --SH, --SC₁ -C₆ alkyl, or --S(CH₂)_(n) NR^(a) R^(b), or R²and R³ together can form a cycloalkyl ring that may contain one or moreheteroatom;

R^(a) and R^(b) are each independently hydrogen or C ₁ -C₆ alkyl, n is 0to 5, and the pharmaceutically acceptable salts, esters, amides, andprodrugs thereof In a preferred embodiment, the protein tyrosine kinaseis SRC. In another preferred embodiment, the protein tyrosine kinase isEGFr. In another preferred embodiment, the protein tyrosine kinase isFGFr. In still another preferred embodiment, the protein tyrosine kinaseis PDGFr.

The present invention also provides the compounds:

1-(3-Hydroxyphenyl)benzimidazole;

1-(3-Aminophenyl)benzimidazole;

1-(3 -Formylphenyl)benzimidazole;

1 -(4-Carbomethoxyphenyl)benzimidazole;

4-Hydroxy-1-phenylbenzimidazole;

5-Methyl-1-phenylbenzimidazole;

1-Phenylbenzimidazole-5-carboxylic acid;

Methyl 1-Phenylbenzimidazole-5-carboxylate;

1-Phenylbenzimidazole-5-carboxamide;

5-Acetyl-1-phenylbenzimidazole;

1-Phenylbenzimidazole-5-carboxaldehyde;

1-Phenylimidazo-1H-imidazo[4,5-c]pyridine;

5-Acetamido-1-phenylbenzimidazole;

5-Propionamido-1-phenylbenzimidazole;

5-Acrylamido-1-phenylbenzimidazole;

5-(Hydroxymethyl)-1-phenylbenzimidazole;

5-Ethoxy-1-phenylbenzimidazole;

1-Phenyl-5-propoxybenzimidazole;

5-Isopropoxy-1-phenylbenzimidazole;

5-Butoxy-1-phenylbenzimidazole

5-Allyloxy-1-phenylbenzimidazole;

5-(2,3-Dihydroxypropoxy)-1-phenylbenzimidazole;

5-(2,3-Epoxypropoxy)-1-phenylbenzimidazole;

5-(2-Aminoethoxy)-1-phenylbenzimidazole;

5-[2-(N,N-Dimethylamino)ethoxy]-1-phenylbenzimidazole;

5-[3-(N,N-Dimethylamino)propoxy]-1-phenylbenzimidazole;

5-[2-(4-Morpholino)ethoxy]-1-phenylbenzimidazole;

5-[3-(4-Morpholino)propoxy]-1-phenylbenzimidazole;

Methyl 1-phenylbenzimidazole-6-carboxylate;

6-Ethoxy-1-phenylbenzimidazole;

1-(4-Aminophenyl)-5-methoxybenzimidazole;

5,6-Dihydroxy-1-phenylbenzimidazole;

5,6-Methylenedioxy-1-phenylbenzimidazole;

5-Methoxy-6-methyl-1-phenylbenzimidazole;

5-Hydroxy-6-methyl-1-phenylbenzimidazole;

Methyl 5-methoxy-1-phenylbenzimidazole-6-carboxylate;

5-Hydroxy-1-phenylbenzimidazole-6-carboxylic acid;

6-Hydroxymethyl-5-methoxy-1-phenylbenzimidazole;

5-Methoxy-1-phenylbenzimidazole-6-carboxaldehyde;

5-Methoxy-1-(2-thienyl)benzimidazole;

5-Methoxy-1-(3-thienyl)benzimidazole;

5-(4-Hydroxybutoxy)-1-phenylbenzimidazole;

5-[4-(N,N-Dimethylamino)butoxy]-1-phenylbenzimidazole;

5-[4-(4-Morpholino)butoxyl]-1]phenylbenzimidazole;

4,5-Dimethoxy-1-phenylbenzimidazole;

4,5-Dihydroxy- 1-phenylbenzimidazole;

5-Hydroxy-4-methoxy-1-phenylbenzimidazole;

4-Hydroxy-5-methoxy-1-phenylbenzimidazole;

4-Bromo-5-hydroxy-1-phenylbenzimidazole;

4-Bromo-5-allyloxy-1-phenylbenzimidazole; or

5-(Methylthio)-1-phenylbenzimidazole.

DETAILED DESCRIPTION OF THE INVENTION

The present invention provides compounds of Formula I that can be usedto treat or prevent cancer, restenosis, atherosclerosis, or psoriasis.

The term "alkyl" means a straight or branched chain hydrocarbon.Representative examples of alkyl groups are methyl, ethyl, propyl,isopropyl, isobutyl, butyl, tert-butyl, sec-butyl, pentyl, and hexyl.

The term "halogen" includes chlorine, fluorine, bromine, and iodine.

The term "alkenyl" means a branched or straight chain hydrocarbon havingone or more carbon-carbon double bond.

The term "aryl" means an aromatic hydrocarbon. Representative examplesof aryl groups include phenyl and naphthyl.

The term "heteroatom" includes oxygen, nitrogen, and sulfur.

The term "heteroaryl" means an aryl group wherein one or more carbonatom of the aromatic hydrocarbon has been replaced with a heteroatom.Examples of heteroaryl radicals include, but are not limited to,pyridyl, imidazolyl, pyrrolyl, thienyl, furyl, pyranyl, pyrimidinyl,pyridazinyl, indolyl, quinolyl, naphthyridinyl, and isoxazolyl.

The term "cycloalkyl" means a cyclic hydrocarbon. Examples of cycloalkylgroups include cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl.

The symbol "-" means a bond.

The term "patient" means all animals including humans. Examples ofpatients include humans, cows, dogs, cats, goats, sheep, and pigs.

The term "substituted" means that the base organic radical has one ormore substituents. For example, substituted aryl means an aryl radicalthat has one or more substituents. Substituents include, but are notlimited to, halogen, C₁ -C₈ alkyl, --CN, CF₃, --NO₂, --NH₂, --NHC₁ -C₈alkyl, --N(C₁ -C₈ alkyl)₂, --OC₁ -C₈ alkyl, and --OH.

The term "heterocycle" means a cycloalkyl group wherein one or morecarbon atom is replaced with a heteroatom. Examples of heterocyclesincludes, but are not limited to, pyrrolidinyl, piperidyl, andpiperazinyl.

Those skilled in the art are easily able to identify patients havingcancer, atherosclerosis, psoriasis, restenosis, or at risk of havingatherosclerosis or restenosis. For example, patients who are at risk ofhaving restenosis include, but are not limited to, patients havingundergone balloon angioplasty or other surgical vascular procedures.

A therapeutically effective amount is an amount of a compound of FormulaI, that when administered to a patient, ameliorates a symptom of thedisease.

The term "cancer" includes, but is not limited to, the followingcancers:

breast;

ovary;

cervix;

prostate;

testis;

esophagus;

glioblastoma;

neuroblastoma;

stomach;

skin, keratoacanthoma;

lung, epidermoid carcinoma, large cell carcinoma, adenocarcinoma;

bone;

colon, adenocarcinoma, adenoma;

pancreas, adenocarcinoma;

thyroid, follicular carcinoma, undifferentiated carcinoma, papillarycarcinoma;

seminoma;

melanoma;

sarcoma;

bladder carcinoma;

liver carcinoma and biliary passages;

kidney carcinoma;

myeloid disorders;

lymphoid disorders, Hodgkin's, hairy cells;

buccal cavity and pharynx (oral), lip, tongue, mouth, pharynx;

small intestine;

colon-rectum, large intestine, rectum;

brain and central nervous system; and

leukemia.

The compounds of the present invention can be administered to a patienteither alone or a part of a pharmaceutical composition. The compositionscan be administered to patients either orally, rectally, parenterally(intravenously, intramuscularly, or subcutaneously), intracisternally,intravaginally, intraperitoneally, intravesically, locally (powders,ointments, or drops), or as a buccal or nasal spray.

Compositions suitable for parenteral injection may comprisephysiologically acceptable sterile aqueous or nonaqueous solutions,dispersions, suspensions or emulsions, and sterile powders forreconstitution into sterile injectable solutions or dispersions.Examples of suitable aqueous and nonaqueous carriers, diluents, solventsor vehicles include water, ethanol, polyols (propyleneglycol,polyethyleneglycol, glycerol, and the like), suitable mixtures thereof,vegetable oils (such as olive oil) and injectable organic esters such asethyl oleate. Proper fluidity can be maintained, for example, by the useof a coating such as lecithin, by the maintenance of the requiredparticle size in the case of dispersions and by the use of surfactants.

These compositions may also contain adjuvants such as preserving,wetting, emulsifying, and dispensing agents. Prevention of the action ofmicroorganisms can be ensured by various antibacterial and antifungalagents, for example, parabens, chlorobutanol, phenol, sorbic acid, andthe like. It may also be desirable to include isotonic agents, forexample sugars, sodium chloride, and the like. Prolonged absorption ofthe injectable pharmaceutical form can be brought about by the use ofagents delaying absorption, for example, aluminum monostearate andgelatin.

Solid dosage forms for oral administration include capsules, tablets,pills, powders, and granules. In such solid dosage forms, the activecompound is admixed with at least one inert customary excipient (orcarrier) such as sodium citrate or dicalcium phosphate or (a) fillers orextenders, as for example, starches, lactose, sucrose, glucose,mannitol, and silicic acid, (b) binders, as for example,carboxymethylcellulose, alignates, gelatin, polyvinylpyrrolidone,sucrose, and acacia, (c) humectants, as for example, glycerol, (d)disintegrating agents, as for example, agar-agar, calcium carbonate,potato or tapioca starch, alginic acid, certain complex silicates, andsodium carbonate, (e) solution retarders, as for example paraffin, (f)absorption accelerators, as for example, quaternary ammonium compounds,(g) wetting agents, as for example, cetyl alcohol, and glycerolmonostearate, (h) adsorbents, as for example, kaolin and bentonite, and(i) lubricants, as for example, talc, calcium stearate, magnesiumstearate, solid polyethylene glycols, sodium lauryl sulfate, or mixturesthereof. In the case of capsules, tablets, and pills, the dosage formsmay also comprise buffering agents.

Solid compositions of a similar type may also be employed as fillers insoft and hard-filled gelatin capsules using such excipients as lactoseor milk sugar as well as high molecular weight polyethyleneglycols, andthe like.

Solid dosage forms such as tablets, drageees, capsules, pills, andgranules can be prepared with coatings and shells, such as entericcoatings and others well known in the art. They may contain opacifyingagents, and can also be of such composition that they release the activecompound or compounds in a certain part of the intestinal tract in adelayed manner. Examples of embedding compositions which can be used arepolymeric substances and waxes. The active compounds can also be inmicro-encapsulated form, if appropriate, with one or more of theabove-mentioned excipients.

Liquid dosage forms for oral administration include pharmaceuticallyacceptable emulsions, solutions, suspensions, syrups, and elixirs. Inaddition to the active compounds, the liquid dosage forms may containinert diluents commonly used in the art, such as water or othersolvents, solubilizing agents and emulsifiers, as for example, ethylalcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzylalcohol, benzyl benzoate, propyleneglycol, 1,3-butyleneglycol,dimethylformamide, oils, in particular, cottonseed oil, groundnut oil,corn germ oil, olive oil, castor oil and sesame oil, glycerol,tetrahydrofurfuryl alcohol, polyethyleneglycols and fatty acid esters ofsorbitan or mixtures of these substances, and the like.

Besides such inert diluents, the composition can also include adjuvants,such as wetting agents, emulsifying and suspending agents, sweetening,flavoring, and perfuming agents.

Suspensions, in addition to the active compounds, may contain suspendingagents, as for example, ethoxylated isostearyl alcohols, polyoxyethylenesorbitol and sorbitan esters, microcrystalline cellulose, aluminummetahydroxide, bentonite, agar-agar and tragacanth, or mixtures of thesesubstances, and the like.

Compositions for rectal administrations are preferably suppositorieswhich can be prepared by mixing the compounds of the present inventionwith suitable non-irritating excipients or carriers such as cocoabutter, polyethyleneglycol or a suppository wax, which are solid atordinary temperatures but liquid at body temperature and therefore, meltin the rectum or vaginal cavity and release the active component.

Dosage forms for topical administration of a compound of this inventioninclude ointments, powders, sprays, and inhalants. The active componentis admixed under sterile conditions with a physiologically acceptablecarrier and any preservatives, buffers, or propellants as may berequired. Ophthalmic formulations, eye ointments, powders, and solutionsare also contemplated as being within the scope of this invention.

The term "pharmaceutically acceptable salts, esters, amides, andprodrugs" as used herein refers to those carboxylate salts, amino acidaddition salts, esters, amides, and prodrugs of the compounds of thepresent invention which are, within the scope of sound medical judgment,suitable for use in contact with the tissues of patients without unduetoxicity, irritation, allergic response, and the like, commensurate witha reasonable benefit/risk ratio, and effective for their intended use,as well as the zwitterionic forms, where possible, of the compounds ofthe invention. The term "salts" refers to the relatively non-toxic,inorganic and organic acid addition salts of compounds of the presentinvention. These salts can be prepared in situ during the finalisolation and purification of the compounds or by separately reactingthe purified compound in its free base form with a suitable organic orinorganic acid and isolating the salt thus formed. Representative saltsinclude the hydrobromide, hydrochloride, sulfate, bisulfate, nitrate,acetate, oxalate, valerate, oleate, palmitate, stearate, laurate,borate, benzoate, lactate, phosphate, tosylate, citrate, maleate,fumarate, succinate, tartrate, naphthylate mesylate, glucoheptonate,lactobionate and laurylsulphonate salts, and the like. These may includecations based on the alkali and alkaline earth metals, such as sodium,lithium, potassium, calcium, magnesium, and the like, as well asnon-toxic ammonium, quaternary ammonium and amine cations including, butnot limited to ammonium, tetramethylammonium, tetraethylammonium,methylamine, dimethylamine, trimethylamine, triethylamine, ethylamine,and the like. (See, for example, S. M. Berge, et al., "PharmaceuticalSalts," J Pharm. Sci., 1977; 66: 1-19 which is incorporated herein byreference.)

Examples of pharmaceutically acceptable, non-toxic esters of thecompounds of this invention include C_(1-C) ₆ alkyl esters wherein thealkyl group is a straight or branched chain. Acceptable esters alsoinclude C₅ -C₇ cycloalkyl esters as well as arylalkyl esters such as,but not limited to benzyl. C₁ -C₄ alkyl esters are preferred. Esters ofthe compounds of the present invention may be prepared according toconventional methods.

Examples of pharmaceutically acceptable, non-toxic amides of thecompounds of this invention include amides derived from ammonia, primaryC₁ -C6alkyl amines and secondary C₁ -C₆ dialkyl amines wherein the alkylgroups are straight or branched chain. In the case of secondary aminesthe amine may also be in the form of a 5- or 6-membered heterocyclecontaining one nitrogen atom. Amides derived from ammonia, C₁ -C₃ alkylprimary amines, and C₁ -C₂ dialkyl secondary amines are preferred.Amides of the compounds of the invention may be prepared according toconventional methods.

The term "prodrug" refers to compounds that are rapidly transformed invivo to yield the parent compound of the above formulae, for example, byhydrolysis in blood. A thorough discussion is provided in T. Higuchi andV. Stella, "Pro-drugs as Novel Delivery Systems," Vol 14 of the A.C.S.Symposium Series, and in Bioreversible Carriers in Drug Design, ed.Edward B. Roche, American Pharmaceutical Association and Pergamon Press,1987, both of which are incorporated herein by reference.

In addition, the compounds of the present invention can exist inunsolvated as well as solvated forms with pharmaceutically acceptablesolvents such as water, ethanol, and the like. In general, the solvatedforms are considered equivalent to the unsolvated forms for the purposesof the present invention.

The compounds of the present invention can exist in differentstereoisomeric forms by virtue of the presence of asymmetric centers inthe compounds. It is contemplated that all stereoisomeric forms of thecompounds, as well as mixtures thereof including racemic mixtures, formpart of this invention.

The compounds of the present invention can be administered to a patientat dosage levels in the range of about 0.1 to about 1,000 mg per day.For a normal human adult having a body weight of about 70 kilograms, adosage in the range of about 0.01 to about 100 mg per kilogram of bodyweight per day is preferable. The specific dosage used, however, canvary. For example, the dosage can depended on a numbers of factorsincluding the requirements of the patient, the severity of the conditionbeing treated, and the pharmacological activity of the compound beingused. The determination of optimum dosages for a particular patient iswell known to those skilled in the art.

In addition, it is intended that the present invention cover compoundsmade either using standard organic synthetic techniques, includingcombinatorial chemistry or by biological methods, such as throughmetabolism.

The examples presented below are intended to illustrate particularembodiments of the invention and are not intended to limit the scope ofthe specification, including the claims, in any way.

EXAMPLES

The compounds of Formula I can be prepared by the processes described inthe following Reaction Schemes: ##STR12##

In Scheme 1, base-catalyzed condensation of substituted2-nitrohalobenzenes (II) with substituted anilines (III), using basessuch as NaOAc, NaHCO₃, K₂ CO₃ or aliphatic amines in solvents such asEtOH or aliphatic amines, at temperatures preferably from 60-130° C.,gives the substituted 2-nitrodiphenylamines (VI). Alternatively,copper-catalyzed condensation of substituted 2-nitroanilines (IV) withsubstituted bromobenzenes (V) under basic conditions, using Na₂ CO₃, K₂CO₃ either with or without a suitable solvent, also gives thesubstituted 2-nitrodiphenylamines (VI). Reduction of the2-nitrodiphenylamines (VI), preferably with Raney nickel or byhydrogenation over a palladium/carbon catalyst in a hydroxylic solvent,preferably methanol, gives the substituted 2-aminodiphenylamines (VII).Cyclization of these using formic acid and a mineral acid (preferablyHCl), or using formamidine acetate in a hydroxylic solvent preferably2-methoxyethanol) under reflux, or using trialkyl orthoformates and ageneral published method (H. J. Schaeffer, C. F. Schwender, J. Med.Chem., 1974; 17: 6-8), gave the desired substituted1-phenylbenzimidazoles. ##STR13##

In Scheme 2, Reaction of substituted benzimidazoles (VII) withsubstitued halobenzenes (IX) and a base such as KH, K₂ CO₃ or NaH in asolvent such as DMF or DMSO at 50-150° C., according to a generalpublished procedure (M. F. Mackay, G. J. Trantino, J. F. K. Wilshire,Aust. J. Chem., 1993; 46: 417-425) gave the desired substituted1-phenylbenzimidazoles. ##STR14##

In Scheme 3, substituted 1-phenylbenzimidazoles of Formula I' bearingsuitable substituent groups R' and R^(1') can be converted to other1-phenylbenzimidazoles of Formula I' by appropriate transformation ofthe substituent groups, by known reactions. A non-exhaustive list ofexamples includes the conversions: OMe→OH (HBr/AcOH); NH₂ →Cl (NaNO₂/CuCl); CN→COOH (90% H₂ SO₄); CH₂ OH→CHO (MnO₂); Me→COOH (KMnO₄);COOH→NH₂ (SOCl₂ /NaN₃ /heat); COOH→COOMe(SOCl₂ /MeOH); COOH→CONH₂ (SOCl₂/NH₃); OH→O alkyl (base/alkyl halide); aryl H→aryl Br(N-bromosuccinimide); OH→SR([a] base/Me₂ NC(S)Cl. [b] heat [c] aqueousKOH [d] base/alkyl halide). ##STR15##

In Scheme 4, condensation of substituted 2-nitroanilines (X) andhaloheterocycles (e.g., bromothiophenes XI) using CuI catalyst and abase (preferably K₂ CO₃) gave, for example, substituted2-nitro-N-(2-thienyl)anilines (XII). Cyclization of these by one of themethods of Scheme 1 (e.g., reduction of the nitro group and reactionwith formamidine acetate) gave the desired substituted1-[(hetero)aromatic]benzimidazoles of Formula (I').

R' and R¹ ' as used in Schemes 1-4 and the Examples have the samemeanings as the corresponding substituents of Formula I.

The following abbreviations are used throughout this patent application:

    ______________________________________                                        Et.sub.2 O     Diethyl ether                                                  EtOAc          Ethyl acetate                                                  AcOH           Acetic acid                                                    Me             Methyl                                                         MeOH           Methanol                                                       mp             melting point                                                  DMF            Dimethylformamide                                              DMSO           Dimethylsulfoxide                                              THF            Tetrahydrofuran                                                MeCN           Acetonitrile                                                   NaOH           Sodium hydroxide                                               DIBAL          Diisobutylaluminum hydride                                     NMR            Nuclear Magnetic Resonance                                     ______________________________________                                    

Example 1

1-Phenylbenzimidazole was prepared by the method of H. Hofinann and J.Spindler, Z. Chem., 1976; 16: 52.

Example 2 1-(3-Hydroxyphenyl)benzimidazole Hydrochloride by the Methodof Scheme 3

A solution of 1-(3'-methoxyphenyl)benzimidazole hydrochloride (0.25 g,1.11 mmol) in 48% HBr in glacial acetic acid (15 mL) was refluxed for 48hours and concentrated to dryness. The residue was partitioned between2N NaOH and Et₂ O, and the aqueous portion was carefully neutralizedwith 2N HCl, extracted with EtOAc, and the extract worked up to give asolid which was chromatographed on silica gel. Elution withEtOAc/petroleum ether (1:1) gave Example 2 (0.22 g, 94%). HCl salt: mp(MeOH/Et₂ O) 119° C.

¹ H NMR (D₂ O): δ9.44 (s, 1 H, H-2), 7.93 (dd, J=8.0, 1.0 Hz, 1 H, H-4),7.74-7.62 (m, 3 H), 7.68-7.63 (m, 1 H), 7.56 (dd, J=8.1, 8.1 Hz, 1 H),7.20 (dd, J=8.4, 1.6 Hz, 1 H), 7.15 (dd, J=8.3, 1.6 Hz, 1 H), 7.10 (dd,J=2.1, 2.1 Hz, 1 H, H-2').

¹³ C NMR: δ159.88 (s), 142.30 (d), 136.82 (s), 134.26 (d), 133.70 (s),133.39 (s), 130.01 (d), 129.87 (d), 120.21 (d), 119.02 (d), 117.68 (d),115.76 (d), 114.34 (d). Analysis calculated for C₁₃ H₁₀ N₂ O.HCI.0.25H₂O requires: C, 62.1; H, 4.6; N, 11.2%. Found: C, 62.3; H, 4.6; N, 11.1%.

1-(3-Methoxyphenyl)benzimidazole hydrochloride was prepared by themethod of Scheme 1 as follows: A mixture of 2-fluoronitrobenzene (II:R'═H, hal=F) (7.47 mL, 0.07 mol), 3-methoxyaniline (III: R¹ '=3-OMe)(7.99 mL, 0.07 mol) and K₂ CO₃ (14.69 g, 0.11 mol) in DMF (60 mL) waswarmed at 125° C. with stirring for 18 hours. After removal of thesolvent under reduced pressure, the residue was partitioned betweenEtOAc and 0.5N HCl, and the EtOAc solution was worked up to give an oil.Excess 2-fluoronitrobenzene was removed by distillation under reducedpressure, and the residue was recrystallized from ethanol to give3'-methoxy-2-nitrodiphenylamine (VI: R'═H, R¹ '═3'-OMe) (1.95 g, 58%):mp (EtOH) 59-60° C.

¹ H NMR (CDCl₃): δ9.45 (br, 1 H, NH), 8.19 (dd, J=8.6, 1.6 Hz, 1 H,H-3), 7.39-7.27 (m, 3 H), 6.87 (dd, J=8.1, 1.9 Hz, 1 H), 6.82-6.75 (m, 3H), 3.82 (s, 3 H, OCH₃).

¹³ C NMR δ160.71 (s), 142.79 (s), 139.89 (s), 135.61 (d), 133.26 (s),130.36 (d), 126.58 (d), 117.56 (d), 116.32 (2d), 111.02 (d), 109.89 (d),55.35 (q). Analysis calculated for C₁₃ H₁₂ N₂ O₃ requires: C, 63.9; H,5.0; N, 11.5%. Found: C, 64.1; H, 4.8; N, 11.5%.

Reduction of 3'-methoxy-2-nitrodiphenylamine, followed by reaction withformamidine acetate, as described for the synthesis of Example 12, gave1-(3-methoxyphenyl)benzimidazole hydrochloride, (77%): mp (MeOH/Et₂ O)119.5° C.

¹ H NMR (D₂ O): δ9.49 (s, 1 H, H-2), 7.95 (d, J=8.2 Hz, 1 H, H-4),7.77-7.72 (m, 1 H), 7.68-7.61 (m, 3 H), 7.25 (dd, J=8.2, 2.3 Hz, 1 H,H-4'), 7.16 (dd, J=2.3, 2.1 Hz, 1 H, H-2'), 3.93 (s, 3 H, OCH₃).

¹³ C NMR: δ162.86 (s), 142.24 (d), 136.66 (s), 134.17 (d), 133.46 (s),133.30 (s), 130.09 (d), 129.92 (d), 119.48 (d), 118.75 (d), 117.75 (d),115.60 (d), 112.91 (d), 58.50 (q). Analysis calculated for C₁₄ H₁₂ N₂O.HCl requires: C, 64.5; H, 5.0; N, 10.7%. Found: C, 64.5; H, 5.5; N,10.8%.

Example 3 1-(3-Aminophenyl)benzimidazole by the Method of Scheme 3

Reduction of 1-(3-nitrophenyl)benzimidazole (prepared by the method ofM. A. Khan and J. B. Polya, J. Chem. Soc. (C), 1970: 85-91) with Raneynickel as for Example 8, followed by chromatography on silica gel,eluting with EtOAc, gave Example 3 (71%) mp (EtOAc/hexane) 93-95° C.

¹ H NMR [(CD₃)₂ SO]; δ5.52 (s, 2 H), 6.66 (dd, J=2.2, 8.2 Hz, 1 H), 6.73(d, J=7.0 Hz, 1 H), 6.80-6.82 (m, 1 H), 7.23 (t, J=8.0 Hz, 1 H),7.27-7.35 (m, 2H) 7.61 (d, J=7.1 Hz, 1 H), 7.76 (d, J=7.4 Hz, 1 H), 8.47(s, 1 H); MS(CI) (m+1)/z 210. Analysis calculate d for C₁₃ H₁₁ N₃requires: C, 74.6; H, 5.3; N, 20.1%. Found: C, 74.8; H, 5.5; N, 19.9%.

Example 4 1-(3-Formylphenyl)benzimidazole Hydrochloride by the Method ofScheme 2

A solution of benzimidazole (VIII: R═H) (1.00 g, 8.47 mmol) and3-fluorobenzaldehyde (IX: R^(1') =3-CHO; halogen=F) (1.08 mL, 10.2 mmol)in DMSO (30 mL) was heated with anhydrous K₂ CO₃ (2.34 g, 16.9 mmol) for24 hours at 100° C. to give, after column chromatography on silica geleluting with EtOAc/hexane (1:1) to EtOAc/hexane (3:1), Example 4 as theHCl salt, mp (EtOAc/MeOH) 196-201° C.

¹ H NMR [(CD₃)₂ SO]: δ10.16 (s, 1 H, CHO), 9.40 (s, 1 H, H-2), 8.30 (t,J=1.6 Hz, 1 H, H-2'), 8.13 (m, 2 H, aromatic), 7.93 (m, 2 H, aromatic),7.78 (m, 1 H, aromatic), 7.55 (m, 2 H, H-5, 6). Analytical calculatedfor C₁₄ H₁₀ N₂ O.HCl.0.5H₂ O requires: C, 62.8; H, 4.5; N, 10.5%. Found:C, 63.0; H, 4.4; N, 10.5%.

Example 5 1-(4-Methoxyphenyl)benzimidazole Hydrochloride by the Methodof Scheme 1

Reduction of 4'-methoxy-2-nitrodiphenylamine (VI, R'═H, R^(1') =4-OCH₃)followed by reaction with formamidine acetate as described for synthesisof Example 12 gave Example 5 as the hydrochloride salt, (76%), mp214-215° C. (MeOH/Et₂ O) (F. Montanari and R. Passerini, Boll. sci.facolta chim. ind. Bologna, 1953; 11: 42-45; CA 48: 6436b record a mp of99° C. for the free base form of Example 5).

¹ H NMR (D₂ O): δ9.47 (s, 1 H, H-2), 7.96 (d, J=8.3 Hz, 1 H, H-4), 7.76(ddd, J=8.3, 8.2, 0.7 Hz, 1 H, H-6), 7.66 (ddd, J=8.4, 8.2, 0.7 Hz, 1 H,H-5), 7.59 (d, J=8.4 Hz, 1 H, H-7), 7.57 (d, J=9.0 Hz, 2 H, H-2', 6'),7.19 (d, J=9.0 Hz, 2 H, H-3',5'), 3.98 (s, 3 H, OCH₃).

¹³ C NMR: δ162.78 (s), 141.97 (d), 133.45 (s), 133.03 (s), 129.98 (d),129.79 (d), 128.47 (s), 128.42 (d), 118.03 (d), 117.62 (d), 115.35 (d),58.44 (q). Analytical calculated for C₁₄ H₁₂ N₂ O.HCl requires: C, 64.5;H, 5.0; N, 10.7%. Found: C, 64.6; H, 5.2; N, 10.8%.

4'-Methoxy-2-nitrodiphenylamine (VI, R'═H, R^(1') =4-OCH₃) was preparedby reaction of 2-fluoronitrobenzene and p-anisidine as described abovefor synthesis of the isomeric 3'-methoxy-2-nitrodiphenylamine (61%), mp88-89° C. (MeOH).

¹ H NMR (CDCl₃): δ9.40 (br, 1 H, NH), 8.18 (dd, J=8.6, 1.5 Hz, 1 H,H-3), 7.32 (ddd, J=8.7, 8.6, 0.9 Hz, 1 H, H-5), 7.19 (d, J=8.8 Hz, 2 H,H-2', 6'), 7.00 (dd, J=8.7, 0.9 Hz, 1 H, H-6), 6.95 (d, J=8.8 Hz, 2 H,H-3', 5'), 6.71 (ddd, J=8.6, 8.6, 0.9 Hz, 1 H, H-4), 3.84 (s, 3 H,OCH₃).

¹³ C NMR: δ157.88 (s), 144.44 (s), 135.67 (d), 132.41 (s), 131.13 (s),127.05 (d), 126.53 (d), 116.71 (d), 115.70 (d), 114.92 (d), 55.48 (q).Analytical calculated for C₁₃ H₁₂ N₂ O₃ requires: C, 63.9; H, 4.9; N,11.5%. Found: C, 63.5; H, 5.0; N, 11.3%.

Example 6 1-(4-Hydroxyphenyl)benzimidazole Hydrochloride by the Methodof Scheme 3

Reaction of 1-(4'-methoxyphenyl)benzimidazole hydrochloride with HBr inglacial acetic acid, as described above for synthesis of Example 2, gaveExample 6 as the hydrochloride salt, (86%), mp 255° C. (MeOH/Et₂ O) (M.Tashiro, T. Itoh and G. Fukata, Synthesis, 1982: 217-219 report a mp of192-195° C. for the free base form of Example 6).

¹ H NMR (D₂ O): δ9.52 (s, 1 H, H-2), 7.99 (d, J=8.4 Hz, 1 H, H-4),7.79-7.71 (m, 3 H, H-5, 6,7), 7.63 (d, J=8.8 Hz, 2 H, H-2',6'), 7.17(d,J=8.8 Hz, 2 H, H-3'5').

¹³ C NMR: δ159.92 (s), 141.98 (d), 133.35 (s), 132.60 (s), 129.30 (d),129.26 (d), 128.60 (d), 127.08 (s), 118.87 (d), 117.09 (d), 114.97 (d).Analytical calculated for C₁₃ H₁₀ N₂ O.HCl requires: C, 63.2; H, 4.5; N,11.4%. Found: C, 62.8; H, 4.5; N, 11.4%.

Example 7 1-(4-Carbomethoxyphenyl)benzimidazole by the Method of Scheme2

Reaction of benzimidazole (VIII: R'═H) and methyl 4-fluorobenzoate (IX:R^(1') =COOMe; halogen=F) using a published procedure [M. F. Mackay, G.J. Trantino, J. F. K. Wilshire, Aust. J. Chem., 1993; 46: 417-425],using DMF as solvent, followed by chromatography on silica gel, elutingwith EtOAc/hexane/CH₂ Cl₂ (5:3:2), gave Example 7 (50%), mp 105-107° C.

¹ H NMR [(CD₃)₂ SO]: δ3.91 (s, 3 H), 7.30-7.40 (m, 2 H), 7.73 (d, J=6.8Hz, 1H), 7.80 (d, J=7.0 Hz, 1 H), 7.89 (d, J=8.7 Hz, 2 H), 8.19 (d,J=8.7 Hz, 2 H), 8.69 (s, 1 H); MS(CI) (m+1)/z 253. Analysis calculatedfor C₁₅ H₁₂ N₂ O₂ requires: C, 71.4; H, 4.8; N, 11.1%. Found: C, 71.8;H, 4.9; N, 11.1%.

Example 8 1-(4-Aminophenyl)benzimidazole Dihydrochloride by the Methodof Scheme 3

Raney nickel (0.5 g) was added to a solution of1-(4-nitrophenyl)-benzimidazole (prepared by the method of M. A. Khanand J. B. Polya, J. Chem. Soc. (C), 1970: 85-91) (1.5 g, 6.3 mmol) inTHF (100 mL) under nitrogen. The nitrogen was exchanged for hydrogen andstirred overnight, then filtered. The filtrate was concentrated, and theresidue was chromatographed on silica gel, eluting with EtOAc/hexane(3:1) followed by EtOAc to give Example 8 as an oil. This was dissolvedin Et₂ O/MeCN (1:1, 200 mL) and treated with HCl gas to give thedihydrochloride salt (1.4 g, 79%), mp 278-281° C. (F. Montanari and R.Passerini, Boll. sci. facolta chim. ind. Bologna, 1953, 11, 42-45; CA48: 6436b record a mp of 116° C. for the free base form of Example 8).

¹ H NMR [(CD₃)₂ SO]: δ7.34 (d, J=8.4 Hz, 2 H), 7.58-7.67 (m, 2H),7.7-7.8 (m 3 H), 7.96 (d, J=7.0 Hz, 1 H), 9.82 (s, 1 H); MS(CI) (m+1)/z210. Analysis calculated for C₁₃ H₁₁ N₃.2HCl.0.25H₂ O requires: C, 54.7;H, 4.8; N, 14.7%. Found: C, 54.5; H, 4.8; N, 14.6%.

Example 9 1-(4-Formylphenyl)benzimidazole by the Method of Scheme 2

Reaction of benzimidazole (VIII: R'═H) with 4-fluorobenzaldehyde (IX:R^(1') =4-CHO; halogen=F) using a published procedure [M. F. Mackay, G.J. Trantino, J. F. K. Wilshire, Aust. J. Chem., 1993; 46: 417-425],using DMF as the solvent, followed by chromatography on silica gel,eluting with EtOAc/hexane/CH₂ Cl₂ (1:4:1 followed by 10:10:1), gaveExample 9 (58%), mp (EtOAc/hexane) 98-99° C. (I. Sircar, B. L. Duell, J.A. Bristol, R. E. Weishaar, and D. B Evans, J. Med. Chem., 1987; 30:1023-1029 report a mp of 100-101° C. for Example 9).

¹ H NMR [(CD₃)₂ SO]: δ7.30-7.40 (m, 2 H), 7.77 (d, J=7.0 Hz, 1 H), 7.82(d, J=7.0 Hz, 1 H), 7.98 (d, J=8.4 Hz, 2 H), 8.16 (d, J=8.7 Hz, 2 H),8.72 (s, 1 H), 10.11 (s, 1 H); MS(CI) (m+1)/z 223. Analysis calculatedfor C₁₄ H₁₀ N₂ O requires: C, 75.7; H, 4.5; N, 12.6%. Found: C, 75.7; H,4.6; N, 12.6%.

Example 10

1-(4-Pyridinyl)benzimidazole dihydrochloride was prepared by the methodof M. A. Khan and J. B. Polya, J Chem. Soc. (C), 1970: 85-91.

Example 11 4-Hydroxy-1-phenylbenzimidazole Hydrochloride by the Methodof Scheme 3

Demethylation of 4-methoxy-1-phenylbenzimidazole hydrochloride with HBrin glacial acetic acid as described for synthesis of Example 2 gaveExample 11 as the HCl salt, (77%): mp (MeOH/Et₂ O) 238-240° C.

¹ H NMR (D₂ O): δ9.41 (s, 1 H, H-2), 7.74-7.67 (m, 5 H, Ph), 7.47(dd,J=8.3, 8.2 Hz, 1 H, H-6), 7.22 (d, J=8.3 Hz, 1 H, H-7), 7.08 (dd,J=8.2 Hz, 1 H, H-5);

¹³ C NMR: δ147.45 (s), 141.70 (d), 136.00 (s), 135.85 (s), 133.32 (d),133.10 (d), 131.01 (d), 127.47 (d), 123.81 (s), 114.17 (d), 106.90 (d).Analytical required for C₁₃ H₁₀ N₂ O.HCl.0.25H₂ O: C, 62.2; H, 4.6; N,11.4%. Found: C, 62.5; H, 4.4; N, 11.2%.

4-Methoxy-1-phenylbenzimidazole hydrochloride was prepared by the methodof Scheme 1 as follows: A suspension of 3-methoxy-2-nitroaniline (IV:R'=3-OMe) (5.00 g, 0.030 mol), K₂ CO₃ (2.21 g, 0.016 mol) and cuprousiodide (50 mg, 0.26 mmol) in bromobenzene (V: R^(1') ═H) (10 mL) wasrefluxed with vigorous stirring for 18 hours, and the excess ofbromobenzene was removed under reduced pressure. The residue waspartitioned between EtOAc and water, filtered through Celite, and theorganic layer worked up and chromatographed on silica gel.EtOAc/petroleum ether (1:19) eluted 3-methoxy-2-nitrodiphenylamine (VI:R'=3-OMe, R^(1') ═H) (4.00 g, 54%): mp (aqueous MeOH) 95° C.

¹ H NMR (CDCl₃): δ7.52 (br, 1 H, NH), 7.33 (dd, J=8.6, 8.3 Hz, 1 H,H-5), 7.23-7.08 (m, 5 H, Ph), 6.84 (dd, J=8.6, 0.8 Hz, 1 H, H-6), 6.44(dd, J=8.3, 0.8 Hz, 1 H, H-4), 3.91 (s, 3 H, OCH₃).

¹³ C NMR: δ154.33 (s), 140.32 (s), 140.06 (s), 132.70 (d), 129.51 (d),123.96 (d), 121.82 (d), 108.86 (d), 102.57 (d), 56.53 (q). Analysiscalculated for C₁₃ H₁₂ N₂ O₃ requires: C, 63.8; H, 5.0; N, 11.5%. Found:C, 63.8; H, 4.9; N, 11.4%.

Reduction of the above nitrodiphenylamine with H₂ /Pd--C followed byreaction of the crude phenylenediamine with formamidine acetate, as forthe synthesis of Example 12, gave 4-methoxy-1-phenylbenzimidazole as theHCl salt (74%): mp 191-193° C.

¹ H NMR (D₂ O): δ9.35 (s, 1 H, H-2), 7.71-7.70 (m, 3 H, Ph), 7.64-7.62(m, 2 H, Ph), 7.53 (dd, J=8.4, 8.1 Hz, 1 H, H-6), 7.24 (d, J=8.4 Hz, 1H, H-7), 7.15 (d, J=8.1 Hz, 1 H, H-5), 4.08 (s, 3 H, OCH₃). ¹³ C NMR:δ150.50 (s), 141.65 (d), 135.98 (s), 135.31 (s), 133.24 (d), 133.11 (d),130.96 (d), 127.28 (d), 124.57 (s), 110.09 (d), 107.48 (d), 59.02 (q).Analysis calculated for C₁₄ H₁₂ N₂ O.HCl requires: C, 64,5; H, 5.0; N,10.7%. Found: C, 64.4; H, 5.0; N, 10.6%.

Example 12 5-Methyl-1-phenylbenzimidazole Hydrochloride by the Method ofScheme 1

A mixture of 4-chloro-3-nitrotoluene (II: R'=4-Me, halogen=Cl) (1.00 mL,7.56 mmol), aniline (III: R^(1') ═H) (6.89 mL, 0.075 mmol) and sodiumacetate (1.24 g, 0.015 mol) was refluxed under nitrogen for 18 hours.The cooled product was partitioned between EtOAc and water, and theorganic portion was washed with 2N HCl, then brine, and worked up togive an oil which was chromatographed on silica gel. Elution withpetroleum ether gave the 4-methyldiphenylamine (VI: R'═4-Me, R^(1') ═H)as an orange oil (1.01 g, 58%) which was used directly. A solution ofthis (1.01 g, 4.42 mmol) in 1:1 MeOH/EtOAc (60 mL) was hydrogenated over5% Pd--C for 3 hours. After removal of the catalyst and concentration todryness under reduced pressure, the residue was dissolved in2-methoxyethanol (100 mL) containing formamidine acetate (0.92 g, 8.84mmol) and the solution refluxed for 3 hours. After removal of thesolvent under reduced pressure, the residue was partitioned betweenEtOAc and water. The organic portion was worked up to give an oil whichwas chromatographed on silica gel. Elution with EtOAc/petroleum ether(1:5) gave foreruns, while EtOAc/petroleum ether (1:1) gave Example 12(0.91 g, 98%). HCl salt: mp (MeOH/Et₂ O) 196-200° C.

¹ H NMR (D₂ O): δ9.40 (s, 1 H, H-2), 7.76-7.72 (m, 3 H, Ph), 7.68 (br s,1 H, H-4),7.63-7.59 (m,2H, Ph),7.50 (d, J=8.6 Hz, 1 H, H-7),7.43 (br d,J=8.6 Hz, 1 H, H-6),2.55 (s, 3 H, CH₃).

¹³ C NMR: δ114.66 (d), 141.08 (s), 135.74 (s), 133.54 (s), 133.28 (d),133.15 (d), 131.68 (s), 131.37 (d), 127.04 (d), 116.93 (d), 115.07 (d),23.38 (q). Analysis calculated for C₁₄ H₁₂ N₂.HCl requires: C, 68.7;H5.4; N, 11.5%. Found: C, 68.9; H, 5.6; N, 11.6%.

Example 13 5-Methoxy-1-phenylbenzimidazole Hydrochloride by the Methodof Scheme 1

A mixture of 4-methoxy-2-nitroaniline (IV, R'═OCH₃) (10.0 g, 59 mmol) K₂CO₃ (16.4 g, 120 mmol), CuI (0.20 g, 1.05 mmol) and bromobenzene (V,R^(1') ═H) (30 mL) was heated under reflux in nitrogen for 48 hours.Excess bromobenzene was removed under reduced pressure, the residue waspartitioned between EtOAc and water, and the mixture was filteredthrough Celite. The organic layer was then worked up, and the residuewas chromatographed on silica gel. Petroleum ether eluted4-methoxy-2-nitrodiphenylamine (VI, R'═4-OCH₃, R^(1') ═H) (8.92 g, 62%),which was used directly. Hydrogenation of this over Pd--C in MeOH, andtreatment of the product with formamidine acetate as described above forsynthesis of Example 12, gave Example 13 as the hydrochloride salt (7.54g, 49% overall): mp (MeOH/Et₂ O) 190-193° C. (Y. S. Tsizin and S. A.Chemyak, Khim. Geterotsikl. Soedin., 1978;12: 1680-1683; CA 90: 137728have reported the synthesis of Example 13).

¹ H NMR (D₂ O): δ9.39 (s, 2 H, H-2), 7.76-7.72 (m, 3 H, Ph), 7.65-7.61(m, 2 H, Ph), 7.55 (d, J=9.2 Hz, 1 H, H-7), 7.37 (d, J=2.3 Hz, 1 H,H-4), 7.19 (dd, J=9.2, 2.3 Hz, 1 H, H-6), 3.95 (s, 3 H, OCH₃);

¹³ C NMR: δ161.43 (s), 141.60 (d), 136.76 (s), 134.58 (s), 133.32 (d),133.16 (d), 128.28 (s), 127.05 (d), 119.74 (d), 116.57 (d), 99.78 (d),58.84 (q). Analysis calculated for C₁₄ H₁₂ N₂ O.HCl requires: C, 64.5;H, 5.0; N, 10.7%. Found: C, 64.3; H, 5.1; N, 10.6%.

Example 14 5-Hydroxy-1-phenylbenzimidazole by the Method of Scheme 3

Demethylation of 5-methoxy- 1-phenylbenzimidazole hydrochloride with HBrin glacial acetic acid, as described above for preparation of Example 2gave Example 14 (83%): mp (EtOAc/hexane) 247-249° C. (K. Fries, Ann.,1927;454: 207 reports a mp of 244° C. for Example 14).

¹ H NMR [(CD₃)₂ SO]: δ9.25 (s, 1 H), 8.43 (s, 1 H), 7.60-7.70 (m, 4 H),7.40-7.50 (m, 2 H), 7.07 (d, J=2.4 Hz, 1 H), 6.82 (dd, J=8.7, 2.4 Hz, 1H). Analysis calculated for C₁₃ H₁₀ N₂ O requires: C, 74.3; H, 4.8; N,13.3%. Found: C, 74.3; H, 5.0; N, 13.2%.

Example 15 5-Chloro-1-phenylbenzimidazole by the Method of Scheme 1

Reaction of 5-chloro-2-nitrodiphenylamine (VI, R'=4-Cl, R^(1') ═H) withRaney nickel in MeOH gave 5-chloro-2-aminodiphenylamine (VII, R═4-Cl,R^(1') ═H) (97%), which was used directly. Reaction of this with formicacid, followed by chromatography of the product on silica gel, elutingwith EtOAc/hexane (1:1), gave Example 15 (71%), mp 95-97° C. (F.Montanari and R. Passerini, Boll. sci. facolta chim. ind. Bologna, 1953;11: 42-45; CA 48: 6436b, record a mp of 118° C. for Example 15).

¹ H NMR [(CD₃)₂ SO]: δ7.35 (dd, J=2.1, 8.6 Hz, 1 H), 7.4-7.5 (m, 1 H),7.5-7.7 (m, 5 H), 7.74 (d, J=8.8 Hz, 1 H), 8.57 (s, 1 H); MS(CI) (m+1)/z229. Analysis calculated for C₁₃ H₉ CIN₂ requires: C, 68.3; H, 4.0; N,12.2%. Found: C, 68.4; H, 4.1; N, 12.3%.

Example 16 1-Phenylbenzimidazole-5-carboxylic Acid Hydrochloride by theMethod of Scheme 3

A solution of the ester Example 17 (1.00 g, 4.20 mmol) and 2N NaOH (10mL) in MeOH (60 mL) was stirred at 20° C. for 1 hour, then concentratedunder reduced pressure to small volume. The residue was diluted withwater, washed with Et₂ O, and carefully neutralized with 3N HCl toprecipitate Example 16 (0.92 g, 97%). HCl salt; mp (MeOH/Et₂ O) 240° C.(dec.).

¹ H NMR [D₂ O/(CD₃)₂ SO]: δ9.66 (s, 1 H, H-2), 8.55 (d, J=0.8 Hz, 1 H,H-4), 8.23 (dd, J=8.7, 0.8 Hz, 1 H, H-6), 7.89 (d, J=8.7 Hz, 1 H, H-7),7.77 (br s, 5 H, Ph).

¹³ C NMR: δ170.37 (s), 144.74 (d), 136.39 (s), 135.09 (s), 133.48 (s),133.06 (d), 132.82 (d), 130.96 (s), 130.10 (d), 126.95 (d), 119.72 (d),115.57 (d). Analysis calculated for C₁₄ H₁₀ N₂ O₂.HCl.1.5H₂ O requires:C, 55.7; H, 4.7; N, 9.3%. Found: C, 55.7; H, 4.1; N, 9.3%.

Example 17 Methyl 1-Phenylbenzimidazole-5-carboxylate by the Method ofScheme 1

A mixture of 4-chloro-3-nitrobenzoic acid (II: R'=4-COOH, halogen=Cl)(5.70 g, 0.023 mol), aniline (III: R^(1') ═H) (3.17 mL, 0.035 mol),N-methylmorpholine (3.24 mnL, 0.025 mol) and copper powder (0.10 g) inisoamyl alcohol (200 mL) was refluxed for 18 hours. The cooled solutionwas filtered through Celite and the filtrate concentrated to drynessunder reduced pressure. The residue was dissolved in EtOAc, washed wellthe 3N HCl, then water, and finally extracted with saturated aqueous Na₂CO₃. Acidification of the extract afforded the 4-carboxydiphenylamine(VI: R=4-COOH, R^(1') ═H) as an orange solid (1.64 g, 28%) which wasdissolved in MeOH (50 mL). The solution was saturated with gaseous HCland then refluxed for 18 hours. After concentration to dryness theresidue was partitioned between EtOAc and water, and the organic portionwas worked up to give crude ester (VI: R'=4-COOMe, R^(1') ═H) as ayellow solid (100%). This material was dissolved in MeOH/EtOAc (1:1) (50mL) and hydrogenated over 5% Pd--C, then treated with formamidineacetate as described above for the synthesis of Example 12. Workup gavean oil which was chromatographed on silica gel. EtOAc/petroleum ether(1:1) gave Example 17 (1.27 g, 23% overall), mp 100-102° C.

¹ H NMR [(CD₃)₂ SO]: δ3.90 (s, 3 H), 7.5-7.6 (m, 1 H), 7.6-7.8 (m, 5 H),7.96 (dd, J=1.5, 8.5 Hz, 1 H), 8.36 (d, J=1.2, 1 H), 8.74 (s, 1 H);MS(CI) (m+1)/z 253. Analysis calculated for C₁₅ H₁₂ N₂ O₂ requires: C,71.4; H, 4.8; N, 11.1%. Found: C, 71.5; H, 4.9; N, 11.0%.

Example 18 1-Phenylbenzimidazole-5-carboxamide Hydrochloride by theMethod of Scheme 3

Reaction of the acid chloride [obtained from the acid Example 16 asdescribed above] with aqueous ammonia gave Example 18 (74%): mp(MeOH/Et₂ O) 225° C. (dec.).

¹ H NMR [D₂ O/(CD₃)₂ SO]: δ9.59 (s, 1 H, H-2), 8.37 (br s, 1 H, H-4),8.04 (dd, J=8.8, 1.4 Hz, 1 H, H-6), 7.87 (d, J=8.8 Hz, 1 H, H-7),7.80-7.72 (m, 5 H, Ph).

¹³ C NMR: δ173.27 (s), 144.99 (d), 136.18 (s), 135.66 (s), 134.36 (s),133.39 (d), 133.22 (d), 128.59 (d), 127.31 (d), 118.08 (d), 116.06 (d).Analysis calculated for C₁₄ H₁₁ N₃ O.HCl.0.25H₂ O requires: C, 60.4; H,4.5; N, 15.1%. Found: C, 60.2; H, 4.5; N, 15.3%.

Example 19

5-Amino-1-phenylbenzimidazole was prepared by the method of M. A.Phillips, J Chem. Soc., 1929: 2820-2828.

Example 20 5-Acetyl-1-phenylbenzimidazole by the Method of Scheme 1

Raney nickel (1.0 g) was added to a solution of4-anilino-3-nitroacetophenone (VI: R'=4-COMe; R^(1') ═H) (2.0 g, 7.8mmol) in THF (100 mL) under nitrogen. The nitrogen was exchanged forhydrogen, and the mixture was stirred for 15.5 hours, then filtered andconcentrated to give crude 4-anilino-3-aminoacetophenone (VII: R=4-COMe;R^(1') ═H) (1.5 g, 87%). Reaction of this with formic acid using a knownprocedure [L. J. Mathias, C. G. Overberger, J. Org. Chem., 1978;43:3518], followed by chromatography of the product on silica gel, elutingwith EtOAc/hexane (1:1), gave Example 20 (68%): mp 106-108° C.

¹ H NMR [(CD₃)₂ SO]: δ2.68 (s, 3 H), 7.55 (t, J=7.2 Hz, 1 H), 7.6-7.8(m, 5 H) 7.95 (dd, J=1.5, 8.5 Hz, 1 H), 8.46 (d, J=1.2, 1 H), 8.74 (s, 1H); MS(CI) (m+1)/z 237. Analysis calculated for C₁₅ H₁₂ N₂ O requires:C, 76.3; H, 5.1; N, 11.9%. Found: C, 76.3; H, 5.0; N, 11.8%.

Example 21 1-Phenylbenzimidazole-5-carboxaldehyde by the Method ofScheme 3

A solution of 5-(hydroxymethyl)-1-phenylbenzimidazole (0.15 g, 0.67mmol) made as described in Example 26 and MnO₂ (0.36 g) in CHCl₃ (10 mL)was stirred overnight, then filtered through Celite and concentrated.The residue was recrystallized from EtOAc/hexane to give Example 21(0.10 g, 70%); mp 114-116° C.

¹ H NMR [(CD₃)₂ SO]: δ7.56 (t, J=7.2 Hz, 1 H), 7.6-7.7 (m, 2 H), 7.7-7.8(m, 4 H), 8.38 (s, 1 H), 8.78 (s, 1 H), 10.11 (s, 1 H); MS(CI) (m+1)/z223. Analysis calculated for C₁₄ H₁₀ N₂ O requires: C, 75.7; H, 4.5; N,12.6%. Found: C, 75.3; H, 4.7; N, 12.5%.

Example 22 1-Phenylimidazo-1H-imidazol[4,5-c]pyridine Hydrochloride bythe Method of Scheme 1

A solution of 4-chloro-3-nitropyridine (II: R'=4-aza, halogen=Cl) (3.22g, 0.020 mol), aniline (III: R^(1') ═H) (1.85 mL, 0.020 mol) and conc.HCl (0.17 mL, 0.02 mol) in 1:1 water/2-methoxyethanol (40 mL) wasrefluxed for 18 hours, then concentrated to dryness. The residue waspartitioned between saturated aqueous NaHCO₃ and EtOAc, and the organicportion was worked up to give an oil which was chromatographed on silicagel. Petroleum ether eluted foreruns, while EtOAc/petroleum ether (1:1)gave 4-(N-phenylamino)-3-nitropyridine (VI: R'=4-aza, R^(1')═H) (2.02 g,41%); mp (EtOAc/petroleum ether) 119° C.

¹ H NMR (CDCl₃): δ9.67 (br s, 1 H, NH), 9.28 (s, 1 H, H-2), 8.25 (dd,J=6.1, 0.8 Hz, 1 H, H-6), 7.51-7.46 (m, 2 H, Ph), 7.38-7.34 (m, 1 H,Ph), 7.31-7.28 (m, 2 H, Ph), 6.94 (d, J=6.1 Hz, 1 H, H-5).

¹³ C NMR: δ153.18 (d), 149.08 (d), 147.53 (s), 136.48 (s), 130.03 (s),130.03 (s), 130.03 (d), 125.36 (d), 109.13 (d). Analysis calculated forC₁₁ H₉ N₃ O₂ requires: C, 61.4; H, 4.2; N, 19.5%. Found: C, 61.2; H,4.1; N, 19.6%.

Reduction of the above nitropyridine, followed by reaction withformamidine acetate as described for synthesis of Example 12, gaveExample 22 (84%). HCl salt: mp (MeOH/Et₂ O) 231-232° C.

¹ H NMR (D₂ O): δ9.43 (s, 1 H, H-2), 9.00 (s, 1 H, H-4), 8.68 (d, J=6.7Hz, 1 H, H-7), 8.20 (d, J=6.7 Hz, 1 H, H-6), 7.76-7.66 (m, 5 H, Ph).

¹³ C NMR: δ153.34 (d), 146.29 (s), 142.53 (s), 137.82 (d), 137.04 (d),136.14 (s), 133.15 (d), 132.76 (d), 127.05 (d), 112.48 (d). Analysiscalculated for C₁₂ H₉ N₃.HCl requires: C, 62.2; H, 4.4; N, 18.4%. Found:C, 62.2; H. 4.5; N, 18.1%.

Example 23 5-Acetamido-1-phenylbenzimidazole Hydrochloride by the Methodof Scheme 3

To a solution of the dihydrochloride salt of the amine Example 19 (0.10g, 0.39 mmol) in water (20 mL) was added acetic anhydride (0.15 mL, 1.59mmol) followed immediately by saturated aqueous sodium acetate solution(5 mL). After vigorous stirring for 15 minutes, the mixture wasextracted with EtOAc and the extract worked up to give the amide Example23. HCl salt (84 mg, 84%): mp (MeOH/Et₂ O) 211-215° C. (hygroscopicpowder).

¹ H NMR (D₂ O): δ9.46 (s, 1 H, H-2), 8.09 (d, J=1.9 Hz, 1 H, H-4),7.77-7.73 (m, 3 H, Ph), 7.66-7.63 (m, 2 H, Ph), 7.61 (d, J=9.1 Hz, 1 H,H-7), 7.44 (dd, J=9.1, 1.9 Hz, 1 H, H-6), 2.15 (s, 3 H, CH₃).

¹³ C NMR: δ175.51 (s), 142.61 (d), 139.34 (s), 135.59 (s), 133.65 (s),133.41 (d), 133.22 (d), 130.37 (s), 126.98 (d), 123.45 (d), 116.27 (d),108.73 (d), 25.81 (q). Analysis calculated for C₁₅ H₁₃ N₃ O.HCl.H₂ Orequires: C, 58.9; H, 5.3; N, 13.7%. Found: C, 58.6; H, 5.3; N, 13.5%.

Example 24 5-Propionamido-1-phenylbenzimidazole Hydrochloride by theMethod of Scheme 3

Reaction of Example 19 with propionic anhydride as described for Example23 gave Example 24 (91%): mp (MeOH/Et₂ O) 218-222° C.

¹ H NMR (D₂ O): δ9.39 (s, 1 H, H-2), 8.13 (d, J=1.8 Hz, 1 H, H-4),7.75-7.66 (m, 6 H, Ph and H-7), 7.49 (dd, J=9.1, 1.8 Hz, 1 H, H-6), 2.47(q, J =7.6 Hz, 2 H, CH₂ CH₃), 1.22 (t, J=7.6 Hz, 3 H, CH₂ CH₃).

¹³ C NMR: δ179.67 (s), 142.99 (d), 139.09 (s), 135.89 (s), 134.33 (s),133.32 (d), 133.17 (d), 131.07 (s), 127.30 (d), 123.87 (d), 116.31 (d),109.69 (d), 32.62 (t), 11.95 (q). Analysis calculated for C₁₆ H₁₅ N₃O.HCl.0.25H₂ O requires: C, 62.7; H, 5.4; N, 13.7%. Found: C, 62.6; H,5.4; N, 13.4%.

Example 25 5-Acrylamido-1-phenylbenzimidazole by the Method of Scheme 3

Acryloyl chloride (0.12 mL, 1.47 mmol) was added dropwise with stirringto a stirred solution of the free base of the amine Example 19 (0.28 g,1.34 mmol) and Et₃ N (0.22 mL, 1.60 mmol) in CH₂ Cl₂ (20 mL). After 30minutes, the solution was washed with saturated NaHCO₃ solution, thenwater, and worked up to give an oil which was chromatographed on silicagel. EtOAc eluted foreruns, followed by Example 25 as a glass (0.10 g,28%): mp (CH₂ Cl₂ /petroleum ether at -20° C.) 180-182° C.

¹ H NMR [(CD₃)₂ SO]: δ10.25 (br s, 1 H, NH), 8.56 (s, 1 H, H-2), 8.27(d, J=1.6 Hz, 1 H, H-4), 7.71-7.64 (m, 4 H, Ph), 7.61 (d, J=8.8 Hz, 1 H,H-7), 7.54 (dd, J=8.8, 1.6 Hz, 1 H, H-6), 7.51-7.48 (m, 1 H, Ph), 6.49(dd, J=16.9, 10.1 Hz, 1 H, CH═CH₂), 6.29 (dd, J=16.9, 1.9 Hz, 1 H,CH═CHH), 5.77 (dd, J=10.1, 1.9 Hz, 1 H, CH═CHH).

¹³ C NMR: δ162.94 (s), 143.90 (s), 143.70 (d), 135.89 (s), 134.29 (s),131.95 (d), 130.00 (d), 129.41 (s), 127.55 (d), 126.47 (t), 123.31 (d),116.41 (d), 100.57 (d), 110.30 (d). Analysis calculated for C₁₆ H₁₃ N₃ Orequires: C, 73.0; H, 5.0; N, 16.0%. Found: C, 72.9; H, 5.2; N, 15.9%.

Example 26 5-(Hydroxymethyl)-1-phenylbenzimidazole by the Method ofScheme 3

A solution of DIBAL (1 M in CH₂ Cl₂ ; 1.3 mL, 1.3 mmol) was added to asolution of methyl 1-phenylbenzimidazole-5-carboxylate (Example 17)(0.15 g, 0.59 mmol) in CH₂ Cl₂ (2 mL) at -78° C. The reaction wasstirred for 1.5 hours, at which time MeOH (10 mL) was added, and thereaction was allowed to warm to room temperature. Sodium tartrate wasadded to dissolve the solids, and the mixture was extracted severaltimes with EtOAc. The combined organic phases were dried (MgSO₄),filtered, and concentrated, and the residue was recrystallized fromEtOAc/hexane to give Example 26 (84 mg, 63%): mp 147-149° C.

¹ H NMR [(CD₃)₂ SO]: δ4.63 (d, J=5.8 Hz, 2 H), 5.24 (t, J=5.8 Hz, 1 H),7.30 (d, J=8.4 Hz, 1 H), 7.4-7.5 (m, 1 H), 7.58 (d, J=8.4 Hz, 1 H),7.6-7.7 (m, 5 H), 8.54 (s, 1 H); MS(CI) (m+1)/z 225. Analysis calculatedfor C₁₄ H₁₂ N₂ O requires: C, 75.0; H, 5.4; N, 12.5%. Found: C, 74.7; H,5.4; N, 12.4%.

Example 27 5-Acetoxy-1-phenylbenzimidazole by the Method of Scheme 1

Reaction of 4-acetoxy-2-aminodiphenylamine (VII: R'=4-OAc, R^(1') ═H)with triethyl orthoformate using a known procedure [H. J. Schaeffer, C.F. Schwender, J. Med. Chem., 1974;17(1): 6-8], followed bychromatography of the product on silica gel, eluting with EtOAc/hexane(1:1), gave Example 27 (77%): mp 92-94° C. (K. Fries, Ann., 1927;454:207 reports a mp of 88° C. for Example 27).

¹ H NMR [(CD₃)₂ SO]: δ2.30 (s, 3 H), 7.10 (dd, J=2.4, 8.9 Hz, 1 H),7.4-7.5 (m, 2 H), 7.6-7.7 (m, 5 H), 8.62 (s, 1 H); MS(CI) (m+1)/z 253.Analysis calculated for C₁₅ H₁₂ N₂ O₂ requires: C, 71.4; H, 4.8; N,11.1%. Found: C, 71.3; H, 4.8; N, 11.0%.

Example 28 5-Ethoxy-1-phenylbenzimidazole Hydrochloride by the Method ofScheme 3

A solution of 5-hydroxy-1-phenylbenzimidazole (Example 14) (0.12 g, 0.57mmol) in 1:1 THF/DMF (5 mL) was added under nitrogen to a stirredsuspension of NaH (16 mg of a 60% dispersion in oil, 0.68 mmol) in THF(5 mL). After 5 minutes, ethyl iodide (55 μL, 0.68 mmol) was added, andthe solution was stirred at 20° C. for 2 hours, then poured into brineand extracted with diethyl ether. The ether extract was washed with 2NNaOH, then water. Evaporation of the organic layer gave a residue thatwas dissolved in EtOAc and percolated through a plug of silica gel togive the ether Example 28 (94 mg, 69%). HCl salt: mp (MeOH/Et₂ O)210-213° C.

¹ H NMR (D₂ O): δ9.37 (s, 1 H, H-2), 7.75-7.71 (m, 3 H, Ph), 7.68-7.65(m, 2 H, Ph), 7.61 (d, J=9.2 Hz, 1 H, H-7), 7.38 (d, J=2.3 Hz, 1 H,H-4), 7.22 (dd, J=9.2, 2.3 Hz, 1 H, H-6), 4.21 (q, J=7.0 Hz, 2 H, OCH₂),1.46 (t, J=7.0 Hz, 3 H, CH₂ CH₃).

¹³ C NMR: δ160.63 (s), 141.72 (d), 135.87 (s), 134.57 (s), 133.34 (d),133.14 (d), 128.55 (s), 127.28 (d), 120.11 (d), 116.73 (d), 100.66 (d),67.98 (t), 16.55 (q). Analysis calculated for C₁₅ H₁₄ N₂ O.HCl requires:C, 65.6; H, 5.5; N, 10.2%. Found: C, 65.7; H, 5.3; N, 10.0%.

Example 29 1-Phenyl-5-propoxybenzimidazole hydrochloride by the Methodof Scheme 3

Reaction of 5-hydroxy-1-phenylbenzimidazole (Example 14) with propyliodide as for Example 28 gave Example 29 (76%); mp (MeOH/Et₂ O) 204-206°C.

¹ H NMR (D₂ O): δ9.38 (s, 1 H, H-2), 7.75-7.71 (m, 3 H, Ph), 7.69-7.65(m, 2 H, Ph), 7.62 (d, J=9.2 Hz, 1 H, H-7), 7.39 (d, J =2.3 Hz, 1 H,H-4), 7.24 (dd, J=9.2, 2.3 Hz, 1 H, H-6), 4.11 (t, J=6.6 Hz, 2 H, OCH₂),1.86 (txq, J=7.5, 6.6 Hz, 2 H, OCH₂ CH₂ CH₃), 1.06 (t, J=7.5 Hz, 3 H,OCH₂ CH₂ CH₃).

¹³ C NMR: δ160.87 (s), 141.69 (d), 135.86 (s), 134.48 (s), 133.36 (d),133.15 (d), 128.53 (s), 127.30 (d), 120.18 (d), 116.75 (d), 100.72 (d),73.86 (t), 24.54 (t), 12.40 (q). Analysis calculated for C₁₆ H₁₆ N₂O.HCl requires: C, 66.5; H, 5.9; N, 9.7%. Found: C, 66.3; H, 5.7; N,9.7%.

Example 30 5-Isopropoxy-1-phenylbenzimidazole Hydrochloride by theMethod of Scheme 3

Reaction of 5-hydroxy-1-phenylbenzimidazole (Example 14) with isopropyliodide as for Example 28 gave Example 30 (66%): mp (MeOH/Et₂ O) 210-211°C.

¹ H NMR (D₂ O): δ9.38 (s, 1 H, H-2), 7.76-7.71 (m, 3 H, Ph), 7.70-7.66(m, 2 H, Ph), 7.63 (d, J=9.2 Hz, 1 H, H-7), 7.44 (d, J=2.3 Hz, 1 H,H-4), 7.24 (dd, J=9.2, 2.3 Hz, 1 H, H-6), 4.79 (sp, J=6.1 Hz, 1 H,OCH(CH₃)₂) 1.41 (d, J=6.1 Hz, 6 H, OCH(CH₃)₂).

¹³ C NMR: δ159.32 (s), 141.93 (d), 135.87 (s), 134.62 (s), 133.32 (d),133.13 (d), 128.74 (s), 127.29 (d), 121.23 (d), 116.80 (d), 102.87 (d),75.51 (d), 23.66 (q). Analysis calculated for C₁₆ H₁₆ N₂ O.HCl requires:C, 66.5; H, 5.9; N, 9.7%. Found: C, 66.2; H, 6.0; N, 9.7%.

Example 31 5-Butoxy-1-phenylbenzimidazole Hydrochloride by the Method ofScheme 3

Reaction of 5-hydroxy-1-phenylbenzimidazole (Example 14) with n-butyliodide as for Example 28 gave Example 31 (72%), mp (MeOH/EtOAc) 196-197°C.

¹ H NMR (D₂ O): δ9.38 (s, 1 H, H-2), 7.75-7.68 (m, 5 H, Ph), 7.65 (d,J=9.2 Hz, 1 H, H-7), 7.41 (d, J=2.2 Hz, 1 H, H-4), 7.26 (dd, J=9.2, 2.2Hz, 1 H, H-6), 4.16 (t, J=6.6 Hz, 2 H, OCH₂ --), 1.82 (m, 2 H, CH₂),1.50 (m, 2 H, CH₂), 0.97 (t, J=7.5 Hz, 3 H, CH₃).

¹³ C NMR: δ160.90 (s), 141.71 (d), 135.89 (s), 134.40 (s), 133.38 (d),133.13 (d), 128.64 (s), 127.42 (d), 120.23 (d), 116.80 (d), 100.74 (d),72.09 (t), 33.11 (t), 21.33 (t), 15.78 (q). Analysis calculated for C₁₇H₁₈ N₂ O.HCl.0.25H₂ O requires: C, 66.4; H, 6.4; N, 9.1%. Found: C,66.8; H, 6.2; N, 9.3%.

Example 32 5-Allyloxy-1-phenylbenzimidazole Hydrochloride by the Methodof Scheme 3

Reaction of 5-hydroxy-1-phenylbenzimidazole (Example 14) with allylbromide as for Example 28 gave Example 32 (88%). HCl salt: mp (MeOH/Et₂O) 200-202° C.

¹ H NMR (D₂ O): δ9.40 (s, 1 H, H-2), 7.76-7.72 (m, 3 H, Ph), 7.67-7.64(m, 2 H, Ph), 7.59 (d, J=9.2 Hz, 1 H, H-7), 7.38 (d, J=2.3 Hz, 1 H,H-4), 7.22 (dd, J=9.2, 2.3 Hz, 1 H, H-6), 6.18-6.11 (m, 1 H, CH═CH₂),5.51 (dd, J17.3, 1.4 Hz, 1 H, CH═CHH), 5.42 (dd, J=10.8, 1.4 Hz, 1 H,CH═CHH), 4.69 (d, J=5.4 Hz, 2 H, OCH₂).

¹³ C NMR: δ160.22 (s), 141.79 (d), 135.77 (s), 135.04 (d), 134.45 (s),133.34 (d), 133.15 (d), 128.53 (s), 127.16 (d), 121.48 (t), 120.19 (d),116.73 (d), 101.08 (d), 72.57 (t). Analysis calculated for C₁₆ H₁₄ N₂O.HCl requires: C, 67.0; H, 5.3; N, 9.8%. Found: C, 67.1; H, 5.5; N,9.8%.

Example 33 5-(2,3-Dihydroxypropoxy)-1-phenylbenzimidazole Hydrochlorideby the Method of Scheme 3

Saturated aqueous KMnO₄ solution was added in portions over 3 hours at20° C. to a stirred solution of 5-allyloxy-1-phenylbenzimidazole(Example 32) (0.25 g, 1.00 mmol) in acetone (50 mL) until no startingmaterial remained. The mixture was filtered through Celite, washing withmore acetone, and the filtrate was concentrated to dryness under reducedpressure. The residue was partitioned between EtOAc and water, and theorganic portion was worked up and chromatographed on silica gel. EtOAceluted foreruns while MeOH/EtOAc (1:19) gave the diol Example 33 (0.18g, 63%). HCl salt: mp (MeOH/Et₂ O) 182-183° C.

¹ H NMR (D₂ O): δ9.35 (s, 1 H, H-2), 7.75-7.68 (m, 5 H, Ph), 7.66 (d,J=9.3 Hz, 1 H, H-7), 7.42 (d, J=2.2 Hz, 1 H, H-4), 7.30 (dd, J=9.3, 2.2Hz, 1 H, H-6 ), 4.27-4.22 (m, 1 H, CHOH), 4.20-4.14 (m, 2 H, ArOCH₂),3.84-3.74 (m, 2 H, CH₂ OH).

¹³ C NMR: δ160.68 (s), 141.95 (d), 135.96 (s), 134.70 (s), 133.31 (d),133.12 (d), 128.92 (s), 127.40 (d), 120.10 (d), 116.80 (d), 100.87 (d),72.70 (d), 72.48 (t), 65.03 (t). Analysis calculated for C₁₆ H₁₆ N₂O₃.HCl.0.5H₂ O requires: C, 58.2; H, 5.5; N, 8.5%. Found: C, 58.0; H,5.0; N, 8.8%.

Example 34 5-(2,3-Epoxypropoxy)-1-phenylbenzimidazole by the Method ofScheme 3

A solution of 5-hydroxy-1-phenylbenzimidazole (Example 14) (0.24 g, 1.14mmol) 1:1 THF/DMF (5 mL) was added under nitrogen to a stirredsuspension of sodium hydride (41 mg of a 60% dispersion in oil, 1.71mmol). After 5 minutes, epichlorohydrin (98 μL, 1.26 mmol) was added,and the solution was refluxed for 4 hours. The cooled solution waspartitioned between diethyl ether and water, and the extract was washedwith 2N NaOH and worked up to give an oil which was chromatographed onsilica gel. EtOAc/petroleum ether (1:1) eluted foreruns, while EtOAceluted the epoxide Example 34 (0.14 g, 46%): mp (EtOAc/petroleum ether)96-97.5° C.

¹ H NMR (CDCl₃): δ8.06 (s, 1 H, H-2), 7.59-7.43 (m, 5 H, Ph), 7.43 (d,J=8.7 Hz, 1 H, H-7), 7.34 (d, J=2.4 Hz, 1 H, H-4), 7.02 (dd, J=8.7, 2.4Hz, 1 H, H-6), 4.32 (dd, J=10.9, 3.1 Hz, 1 H, ArOCHH), 4.04 (dd, J=10.9,5.7 Hz, 1 H, ArOCHH), 3.44-3.40 (m, 1 H, CHO), 2.94 (dd, J=4.9, 4.3 Hz,1 H, CHHO), 2.81 (dd, J=4.9, 2.6 Hz, 1 H, CHHO).

¹³ C NMR: δ155.29 (s), 144.84 (s), 142.51 (d), 136.38 (s), 130.04 (d),128.64 (s), 127.94 (d), 123.79 (d), 114.28 (d), 110.99 (d), 103.67 (d),69.44 (t), 50.17 (d), 44.77 (t). Analysis calculated for C₁₅ H₁₄ N₂ O₂requires: C, 72.2; H, 5.3; N, 10.5%. Found: C, 71.8; H, 4.8; N, 10.6%.

Example 35 5-(2-Aminoethoxy)-1-phenylbenzimidazole Dihydrochloride bythe Method of Scheme 3

A solution of 5-hydroxy-1-phenylbenzimidazole (Example 14) (0.10 g, 0.47mmol) in 1:1 THF/DMF (2 mL) was added under nitrogen to a stirredsuspension of sodium hydride (28.5 mg of a 60% dispersion in oil, 1.19mmol) in THF (4 mL). After 10 minutes, solid 2-bromoethylaminehydrobromide (97 mg, 0.47 mmol) was added, and the mixture was warmed at45° C. for 30 minutes. A further portion of sodium hydride (28.5 mg)followed by bromoethylamine hydrobromide (97 mg) was added and stirringwas continued at this temperature for 1 hour. Water was added, followedby brine, and the mixture was extracted into EtOAc. The organic solutionwas extracted into 2N HCl, and the extract was basified with 2N NaOH,extracted into ether, and the ether solution worked up to give an oilwhich was chromatographed on alumina. EtOAc eluted foreruns whileMeOH/EtOAc (1:9) gave the product Example 35 as a solid (68.5 mg, 58%).DiHCl salt: mp (MeOH/Et₂ O) 252-254° C.

¹ H NMR (D₂ O): δ9.43 (s, 1 H, H-2), 7.78-7.72 (m, 6 H, Ph and H-7),7.51 (d, J=2.3 Hz, 1 H, H-4), 7.38 (dd, J=9.2, 2.3 Hz, 1 H, H-6), 4.46(t, J=4.9 Hz, 2 H, CH₂ NH₃), 3.56 (t, J=4.9 Hz, 2 H, OCH₂).

¹³ C NMR: δ160.02 (s), 142.22 (d), 135.96 (s), 134.61 (s), 133.40 (d),133.16 (d), 129.29 (s), 127.54 (d), 120.04 (d), 117.02 (d), 100.97 (d),67.55 (t), 41.64 (t). Analysis calculated for C₁₅ H₁₅ N₃ O.2HCl.0.5H₂ Orequires: C, 53.7; H, 5.4; N, 12.5%. Found: C, 53.8; H, 5.5; N, 12.5%.

Example 36 5-[2-(N,N-Dimethylamino)ethoxy]-1-phenylbenzimidazoleDihydrochloride by the Method of Scheme 3

Similar reaction of 5-hydroxy-1-phenylbenzimidazole (Example 14) withsodium hydride followed by 2-dimethylaminoethyl chloride hydrochloridegave Example 36 (47%). DiHCl salt: mp (MeOH/Et₂ O) 205-207° C.(hygroscopic foam).

¹ H NMR (D₂ O): δ9.45 (s, 1 H, H-2), 7.77-7.72 (m, 6 H, Ph and H-7),7.53 (d, J=2.3 Hz, 1 H, H-4),7.39 (dd, J=9.2, 2.3 Hz, 1 H, H-6),4.55 (t,J=5.0 Hz, 2 H, CH₂ N⁺ Me₂), 3.73 (t, J=5.0 Hz, 2 H, OCH₂). 3.06 (s, 6 H,N⁺ Me₂).

¹³ C NMR: δ159.71 (s), 142.27 (d), 135.91 (s), 134.42 (s), 133.43 (d),133.15 (d), 129.39 (s), 127.57 (d), 119.97 (d), 117.08 (d), 100.98 (d),65.16 (t), 58.91 (t), 45.63 (q). Analysis calculated for C₁₇ H₁₉ N₃O.2HCl requires: C, 57.5; H, 6.0; N, 11.9%. Found: C, 57.4; H, 6.1; N,11.8%.

Example 37 5-[3--(N,N-Dimethylamino)propoxyl]-1-phenylbenzimidazoleDihydrochloride by the Method of Scheme 3

Similar reaction of 5-hydroxy-1-phenylbenzimidazole (Example 14) withsodium hydride followed by 3-dimethylaminopropyl chloride hydrochloridegave Example 37 (52%). DiHCl salt: mp (MeOH/Et₂ O) dec. above 76° C.(hygroscopic).

¹ H NMR (D₂ O): δ9.41 (s, 1 H, H-2), 7.75-7.71 (m, 6 H, Ph and H-7),7.46 (d, J=2.3 Hz, 1 H, H-4), 7.33 (dd, J=9.2, 2.3 Hz, 1 H, H-6), 4.30(t, J=5.7 Hz, 2 H, CH₂ N⁺ Me₂), 3.44 (t, J=7.7 Hz, 2 H, CH₂ O), 2.98 (s,6 H, N⁺ Me₂), 2.36-2.29 (m, 2 H, CH₂ CH₂ CH₂).

¹³ C NMR: δ160.49 (s), 142.01 (d), 135.94 (s), 134.43 (s), 133.42 (d),133.14 (d), 129.08 (s), 127.58 (d), 120.15 (d), 116.97 (d), 100.76 (d),68.73 (t), 58.21 (t), 45.63 (q), 26.69 (t). Analysis calculated for C₁₈H₂₁ N₃ O.2HCl.3.5H₂ O requires: C, 50.1; H, 7.0; N, 9.7%. Found: C,50.2; H, 7.2; N, 9.8%.

Example 38 5-[2-(4-Morpholino)ethoxy]-1-phenylbenzimidazoleDihydrochloride by the Method of Scheme 3

Similar reaction of 5-hydroxy-1-phenylbenzimidazole (Example 14) withsodium hydride followed by 4-(2-chloroethyl)morpholine hydrochloridegave Example 38 (57%). DiHCl salt: mp (MeOH/Et₂ O) 232-234° C.(hygroscopic foam).

¹ H NMR (D₂ O): δ9.44 (s, 1 H, H-2), 7.78-7.72 (m, 6 H, Ph and H-7),7.53 (d, J=2.3 Hz, 1 H, H-4), 7.39 (dd, J=9.2, 2.3 Hz, 1 H, H-6), 4.60(t, J4.8 Hz, 2 H, CH₂ N⁺), 4.19-3.95 (2×br, 4 H, morpholino CH₂ N⁺),3.79 (t, J=4.8 Hz, 2 H, CH₂ O).

¹³ C NMR: δ159.61 (s), 142.34 (d), 135.95 (s), 134.60 (s), 133.40 (d),133.16 (d), 129.42 (s), 127.55 (d), 119.93 (d), 117.07 (d), 101.05 (d),66.38 (t), 64.70 (t), 58.57 (t), 54.73 (t). Analysis calculated for C₁₉H₂₁ N₃ O₂.2HCl.0.5H₂ O requires: C, 56.3; H, 6.0, N, 10.4%. Found: C,56.4; H, 6.2; N, 10.2%.

Example 39 5-[3-(4-Morpholino)propoxy]-1-phenylbenzimidazoleDihydrochloride by the Method of Scheme 3

Similar reaction of 5-hydroxy-1-phenylbenzimidazole (Example 14) withsodium hydride followed by 4-(3-chloropropyl)morpholine hydrochloridegave Example 39 (52%). DiHCl salt: mp (MeOH/Et₂ O) dec. above 84° C.(hygroscopic solid).

¹ H NMR (D₂ O): δ9.42 (s, 1 H, H-2), 7.81-7.74 (m, 5 H, Ph), 7.72 (d,J=9.2 Hz, 1 H, H-7), 7.47 (d, J=2.2 Hz, 1 H, H-4), 7.34 (dd, J=9.2, 2.2Hz, 1 H, H-6), 4.33 (t, J=5.6 Hz, 2 H, CH₂ N⁺), 4.23 (br d, J=12.9 Hz, 2H, morpholino CH₂ N⁺), 3.92 (br t, J=12.9 Hz, 2 H, morpholino CH₂ N⁺),3.70 (br d, J=12.6 Hz, 2 H, morpholino CH₂ O), 3.52 (t, J=5.6 Hz, 2 H,CH₂ O), 3.36-3.28 (m, 2 H, morpholino CH₂ O), 2.42-2.36 (m, 2 H, CH₂ CH₂CH₂).

¹³ C NMR: δ160.39 (s), 142.00 (d), 135.93 (s), 134.61 (s), 133.35 (d),133.14 (d), 128.94 (s), 127.43 (d), 120.08 (d), 116.89 (d), 100.79 (d),68.65 (t), 66.58 (t), 57.58 (t), 54.62 (t), 25.86 (t). Analysiscalculated for C₂₀ H₂₃ N₃ O₂.2HCl.3.5H₂ O requires: C, 50.7; H, 6.7; N,8.9%. Found: C, 51.1; H, 7.0; N, 8.8%.

Example 40 6-Methoxy-1-phenylbenzimidazole by the Method of Scheme 1

Sodium methoxide (4.4 mL of a 4 M solution in MeOH) was added to asolution of 5-chloro-2-nitrodiphenylamine (VI, R'=5-Cl, R¹ '═H) (2.0 g,8.0 mmol) in MeOH (25 mL). The reaction was heated at reflux for 48hours and cooled to room temperature, and the precipitate of5-methoxy-2-nitrodiphenylamine (VI, R'=5-OCH₃, R¹ '═H) was collected byfiltration, washed with water and dried (1.9 g, 99%); mp (MeOH) 108-110°C.

¹ H NMR [(CD₃)₂ SO]: δ3.73 (s, 3 H), 6.49 (dd, J=2.4, 9.6 Hz, 1 H), 6.53(d, J=2.4Hz, 1 H), 7.24 (t, J=7.2Hz, 1 H), 7.3-7.5 (m, 4 H), 8.13 (d,J=9.6 Hz, 1 H), 9.65 (s, 1 H); MS(CI) (m+1)/z245. Analysis calculatedfor C₁₃ H₁₂ N₂ O₃ requires: C, 63.9; H, 4.9; N, 11.5%. Found: C, 63.8;H, 4.9; N, 11.4%.

A solution of this (1.5 g, 6.15 mmol) in MeOH/THF (1:1, 100 mL) washydrogenated over 5% Pd--C for 15 hours, then filtered and concentrated.The residue was chromatographed on silica gel, eluting with EtOAc/hexane(1:2), to give 5-methoxy-2-aminodiphenylamine (1.05 g, 79%), which wasused directly. Reaction with formic acid followed by chromatography onsilica gel and elution with EtOAc/hexane (1:1), gave Example 40 (73%);mp 71-73° C. (Y. S. Tsizin and S. A. Chemyak, Khim. Geterotsikl.Soedin., 1978;12: 1680-1683; CA 90: 137728 have reported the synthesisof Example 40).

¹ H NMR [(CD₃)₂ SO]: δ3.80 (s, 3 H), 6.93 (dd, J=2.4, 8.7 Hz, 1 H), 7.06d, J=2.4 Hz, 1 H), 7.45-7.55 (m, 1 H), 7.6-7.7 (m, 5 H), 8.41 (s, 1 H);MS(CI) (m+1)/z225. Analysis calculated for C₁₄ H₁₂ N₂ O requires: C,75.0; H, 5.4; N, 12.5%. Found: C, 74.9; H, 5.5; N, 12.4%.

Example 41 6-Hydroxy-1-phenylbenzimidazole Hydrochloride by the Methodof Scheme 3

Reaction of the methyl ether (Example 40) with HBr in glacial aceticacid, as described above for the ether Example 2 gave the phenol(Example 41) as the hydrochloride salt (88%): mp 210-212° C. (Y. S.Tsizin and S. A. Chernyak, Khim. Geterotsikl. Soedin., 1978;12:1680-1683; CA 90: 137728 have reported the synthesis of Example 41).

¹ H NMR (D₂ O): δ9.34 (s, 1 H, H-2), 7.74-7.70 (m, 4 H, Ph and H-4),7.61-7.58 (m, 2 H, Ph), 7.16 (dd, J=9.1, 2.2 Hz, 1 H, H-5), 6.92 (d,J=2.2 Hz, 1 H, H-7).

¹³ C NMR: δ158.22 (s), 141.50 (d), 135.67 (s), 134.77 (s), 133.20 (d),133.10 (d), 127.25 (s), 127.03 (d), 119.65 (d), 118.54 (d), 100.22 (d).

Example 42

6-Chloro-1-phenylbenzimidazole was prepared by the method of F.Montanari and R. Passerini, Boll. sci. facolta chim. ind. Bologna,1953;11: 42-45; CA 48: 6436b.

Example 43 Methyl 1-phenylbenzimidazole-6-carboxylate Hydrochloride bythe Method of Scheme 3

A mixture of 1-phenylbenzimidazole-6-carboxylic acid hydrochloride (0.50g, 2.10 mmol) and SOCl₂ (10 mL) in 1,2-dichloroethane (50 mL) containingDMF (1 drop) was refluxed for 2 hours. The solution was concentrated todryness under reduced pressure. The resulting crude acid chloride wasdissolved in methanol (20 mL) and the solution refluxed for 15 minutes.The methanol was removed under reduced pressure and the residuepartitioned between EtOAc and saturated aqueous NaHCO₃. The organicsolution was worked up to give Example 43 (0.48 g, 91%) as thehydrochloride salt: mp (MeOH/Et₂ O) 200-203° C.

¹ H NMR (D₂ O): δ9.68 (s, 1 H, H-2), 8.35 (br s, 1 H, H-7), 8.30 (br d,J=8.7 Hz, 1 H, H-5), 8.02 (d, J=8.7 Hz, 1 H, H-4), 7.74 (br s, 5 H, Ph),3.95 (s, 3 H, COOCH₃).

¹³ C NMR: δ170.47 (s), 145.03 (d), 136.46 (s), 135.40 (s), 134.11 (s),133.75 (d), 133.25 (d), 131.31 (s), 130.74 (d), 127.72 (d), 118.03 (d),118.01 (d), 55.79 (q). Analysis calculated for C₁₅ H₁₂ N₂ O₂.HClrequires: C, 62.4; H, 4.5; N, 9.7%. Found: C, 62.6; H, 4.4; N, 9.8%.

Example 44 6-Ethoxy-1-phenylbenzimidazole by the Method of Scheme 1

Sodium ethoxide (17.7 mL of a 1 M solution) was added to a solution of5-chloro-2-nitrodiphenylamine (VI: R'=5-Cl, R^(1') ═H) (2.0 g, 8.0 mmol)in MeOH (5 mL). The reaction was heated at reflux for 48 hours, then thereaction was cooled to room temperature and filtered to collect5-ethoxy-2-nitrodiphenylamine (VI: R'=5-OEt, R^(1') ═H) (1.0 g, 50%): mp(EtOH) 101-102° C.

¹ H NMR [(CD₃)₂ SO]: δ1.27 (t, J=7.0 Hz, 3 H), 3.98 (q, J=7.0 Hz, 2 H),6.4-6.5 (m, 2 H), 7.24 (t, J=7.2 Hz, 1 H), 7.3-7.5 (m, 4 H), 8.1-8.2 (m,1 H), 9.63 (s, 1 H); MS(CI) (m+1)/z259. Analysis calculated for C₁₄ H₁₄N₂ O₃ requires: C, 65.1; H, 5.5; N, 10.9%. Found: C, 64.9; H, 5.5; N,10.9%.

Reduction of the 5-ethoxydiphenylamine with Raney nickel as above, usingEtOH/THF (1:1), gave 5-ethoxy-2-aminodiphenylamine (VII: R'=5-OEt, R¹'═H) (92%), which was used directly.

¹ H NMR [(CD₃)₂ SO]: δ1.25 (t, J=7.0 Hz, 3 H), 3.84 (q, J=7.0 Hz, 2 H),4.32 (s, 2 H), 6.43 (dd, J=2.7, 8.5 Hz, 1 H), 6.60 (d, J=2.7 Hz, 1 H),6.65 (d, J=8.5 Hz, 1 H), 6.69 (t, J=7.23 Hz, 1 H), 6.78 (d, J=7.7 Hz, 2H), 7.1-7.2 (m, 3 H); MS(CI) (m+1)/z229.

Reaction of the above aminodiphenylamine with formic acid, followed bychromatography of the product on silica gel, eluting with EtOAc/hexane(1:1), gave Example 44 (82%): mp 66-69° C. (Jacobson, W. Fischer, Chem.Ber., 1892;25: 1000 report a mp of 77-78° C. for Example 44).

¹ H NMR [(CD₃)₂ SO]: δ1.33 (t, J=7.0 Hz, 3 H), 4.04 (d, J=7.0 Hz, 2 H),6.92 (dd, J=2.4, 8.9 Hz, 1 H), 7.04 (d, J=2.2 Hz, 1 H), 7.4-7.5 (m, 1H),7.6-7.7 (m, 5 H), 8.41 (s, 1 H); MS(CI) (m+1)/z239. Analysis calculatedfor C₁₅ H₁₄ N₂ O requires: C, 75.6; H, 5.9; N, 11.8%. Found: C, 75.7; H,6.0; N, 11.7%.

Example 45 1-(4-Aminophenyl)-5-methoxybenzimidazole Dihydrochloride bythe Method of Scheme 1

A mixture of 4-methoxy-2-nitroaniline (IV: R'=4-OMe) (1.00 g, 5.95mmol), K₂ CO₃ (0.34 g, 2.46 mmol), Cul (50 mg) and 4-nitrobromobenzene(V: R^(1') =4-NO₂) (3.00 g, 0.015 mol) was heated with stirring to 150°C. and held at this temperature for 48 hours. The cooled mixture waspartitioned between EtOAc and brine, and the organic portion was workedup and columned on silica gel. EtOAc/petroleum ether (1:1) eluted thediphenylamine (VI: R'=4-OMe, R^(1') =4-NO₂) and a little of the startingmethoxynitroaniline as an inseparable mixture (1.12 g) which was useddirectly. The mixture was dissolved in EtOAc/MeOH (1:1) (50 mL) andhydrogenated over 5% Pd--C (0.20 g) for 3 hours. After removal of thecatalyst by filtration through Celite, the solvent was removed underreduced pressure, and the residue was dissolved in 4N HCl (30 mL)containing formic acid (1 mL) and the solution refluxed for 90 minutes.After concentration to dryness, the residue was basified with 2N NH₄ OHsolution and extracted into EtOAc. Workup gave an oily solid which waschromatographed on silica gel. EtOAc/petroleum ether (1:1) elutedforeruns while EtOAc gave (Example 45) (0.26 g, 17%). DiHCl salt: mp(MeOH/Et₂ O) 237-239° C.

¹ H NMR (D₂ O): δ9.44 (s, 1 H, H-2), 7.86 (d, J=8.7 Hz, 2 H, H-3'5'),7.70 (d, J=9.2 Hz, 1 H, 7), 7.69 (d, J=8.7Hz, 2 H, H-2', 6'), 7.46 (d,J=2.3 Hz, 1 H, H-4), 7.33 (dd, J=9.2, 2.3 Hz, 1 H, H-6), 3.98 (s, 3 H,OCH₃).

¹³ C NMR δ161.69 (s), 142.02 (d), 137.08 (s), 135.28 (s), 134.53 (s),129.41 (d), 128.71 (s), 126.89 (d), 120.08 (d), 116.64 (d), 99.97 (d),58.90 (q). Analysis calculated for C₁₄ H₁₃ N₃ O.2HCl.0.5H₂ O requires:C, 52.3; H, 5.0; N, 13.1%. Found: C, 52.0; H, 5.0; N, 12.6%.

Example 46 5,6-Dimethoxy-1-phenylbenzimidazole Hydrochloride by theMethod of Scheme 1

A mixture of 1-bromo-4,5-dimethoxy-2-nitrobenzene (II: R'4,5-diOMe,halogen=Br) (1.00 g, 4.34 mmol), K₂ CO₃ (1.20 g, 8.70 mmol), Cul (100mg) and aniline (III: R^(1') ═H) (5 mL) was refluxed under nitrogen for5 hours. After diluting with excess 3N HCl, the mixture was extractedwith EtOAc and worked up to give an oil which was chromatographed onsilica gel. EtOAc/petroleum ether (1:5) eluted4,5-dimethoxy-2-nitrodiphenylamine (VI: R'=4,5-diOMe, R^(1') ═H) (0.63g, 60%): mp (MeOH) 120-122° C.

¹ H NMR: δ(CDCl₂) 9.87 (br, 1 H, NH), 7.64 (s, 1 H, H-6), 7.45-7.42 (m.2 H, Ph), 7.32-7.22 (m, 3 H, Ph), 6.63 (s, 1 H, H-3), 3.90, 3.78 (2s, 6H, OCH₃). Analysis required for C₁₄ H₁₄ N₂ O₄.0.5H₂ O requires: C, 59.4;H, 5.3; N, 9.9%. Found: C, 59.8; H, 5.3; N, 9.8%.

Hydrogenation of the above diphenylamine followed by reaction withformamidine acetate gave Example 46 (79%). HCl salt: mp (MeOH/Et2O)217-218.5° C. (P. Jacobson, M. Jaenicke and F. Meyer, Ber., 1896;29:2680-2690 report a mp of 106-107° C. for the free base form of Example46).

¹ H NMR (D₂ O): δ9.26 (s, 1 H, H-2), 7.77-7.72 (m, 3 H, Ph), 7.69-7.64(m, 2 H, Ph), 7.37 (s, 1 H, H-4), 7.07 (s, 1 H, H-7), 3.95-3.83 (2s, 6H, OCH₃).

¹³ CNMR: δ152.13 (2s), 140.18 (d), 135.95 (s), 133.39 (d), 133.23 (d),128.31 (s), 127.66 (s), 127.42 (d), 99.13 (d), 97.27 (d), 59.00 (q),58.91 (q). Analysis calculated for C₁₅ H₁₄ N₂ O₂.HCl requires: C, 62.0;H, 5.2; N, 9.6%. Found: C, 61.4; H, 5.3; N, 9.5%.

Example 47 5,6-Dihydroxy-1-phenylbenzimidazole Hydrochloride by theMethod of Scheme 3

A mixture of the free base of Example 46 (0.80 g, 3.15 mmol) and freshlyprepared pyridine hydrochloride (20 g) was warmed to 220° C. under astream of nitrogen. After 1 hour at this temperature, the cooled mixturewas diluted with water, basified with 2N NaOH, and washed well withdiethyl ether. The aqueous portion was carefully neutralized with 3NHCl, extracted with EtOAc, and worked up to give an oil which waschromatographed on silica gel. EtOAc eluted foreruns while MeOH/EtOAc(1:15) gave Example 47 (0.54 g, 76%). HCl salt: mp (MeOH/Et₂ O) 239-242°C.

¹ H NMR (D₂ O): δ9.16 (s, 1 H, H-2), 7.73-7.68 (m, 3 H, Ph), 7.66-7.62(m, 2 H, Ph), 7.27 (s, 1 H, H-4), 7.08 (s, 1 H, H-7).

¹³ C NMR: δ148.46 (s), 148.43 (s), 140.09 (d), 136.12 (s), 133.17 (d),133.10 (d), 128.35 (s), 127.54 (s), 127.34 (d), 102.43 (d), 100.83 (d).Analysis calculated for C₁₃ H₁₀ N₂ O₂.HCl.1.5H₂ O requires: C, 53.9; H,4.9; N, 9.7%. Found: C, 53.8; H, 4.6; N, 9.5%.

Example 48 5,6-Methylenedioxy-1-phenylbenzimidazole Hydrochloride by theMethod of Scheme 3

A mixture of the benzimidazole Example 47 (0.20 g, 0.88 mmol), 50%aqueous NaOH solution (8 mL), tetrabutylammonium bromide (40 mg, 0.12mmol) and dibromomethane (8 mL) was refluxed for 3 hours. After dilutionwith CH₂ Cl₂, the organic portion was worked up to give an oil which waschromatographed on silica gel. EtOAc/petroleum ether (1:1) elutedExample 48 (0.16 g, 75%). HCl salt: mp (MeOH/Et₂ O) 239-241° C.

¹ H NMR (D₂ O): δ9.21 (s, 1 H, H-2), 7.73-7.69 (m, 3 H, Ph), 7.67-7.63(m, 2 H, Ph), 7.30 (s, 1 H, H-4), 7.14 (s, 1 H, H-7), 6.15 (s, 2 H, OCH₂O).

¹³ C NMR: δ151.12 (s), 151.03 (s), 140.30 (d), 135.97 (s), 133.35 (d),133.12 (d), 129.61 (s), 128.69 (s), 127.55 (d), 105.63 (t), 96.94 (d),95.53 (d). Analysis calculated for C ₁₄ H₁₀ N₂ O₂.HCl.0.25H₂ O requires:C, 60.2; H, 4.2; N, 10.0%. Found: C, 60.0; H, 4.2; N, 10.1%.

Example 49 5-Methoxy-6-methyl-1-phenylbenzimidazole Hydrochloride by theMethod of Scheme 1

A mixture of 4-methoxy-5-methyl-2-nitroaniline (IV: R'=4-OMe, 5-Me)(1.00 g, 5.49 mmol), K₂ CO₃ (0.50 g, 3.62 mmol), Cul (100 mg) andbromobenzene (V: R^(1') ═H) (5 mL) was refluxed under nitrogen withstirring for 4 days. The mixture was concentrated to dryness underreduced pressure, and the residue was partitioned between EtOAc andwater and filtered through Celite. The organic layer was worked up togive an oil which was chromatographed on silica gel. Petroleum ethereluted foreruns, while EtOAc/petroleum ether (1:9) gave4-methoxy-5-methyl-2-nitrodiphenylamine (VI: R'=4-OMe, 5-Me, R^(1') ═H)(1.26 g, 88%): mp (MeOH at -20° C.) 94-96° C.

¹ H NMR (CDCl₃): δ9.46 (brs, 1 H, NH), 7.55 (s, 1 H, H-3), 7.42-7.38 (m,2 H, Ph), 7.26-7.24 (m, 2 H, Ph), 7.22-7.18 (m, 1 H, Ph), 6.98 (d, J=0.7Hz, 1 H, H-6), 3.84 (s, 3 H, OCH₃), 2.17 (s, 3 H, CH₃).

¹³ C NMR: δ149.95 (s), 139.34 (s), 138.78 (s), 138.04 (s), 129.65 (d),124.99 (d), 123.71 (d), 117.70 (d), 104.92(d), 55.77 (q), 17.0 5 (q).Analysis calculated for C₁₄ H₁₄ N₂ O₃ requires: C, 65.1; H, 5.5; N,10.8%. Found: C, 65.1; H, 5.4; N, 10.8%.

Hydrogenation of the above diphenylamine, followed by reaction withformamidine acetate, gave Example 49 (7(1%) HCl salt: mp (MeOH/Et₂ O)214-2175C.

¹ H NMR (D₂ O): δ9.26 (s, 1 H, H-2), 7.73-7.70 (m, 3 H, Ph), 7.63-7.60(m Ph), 7.45 (s, 1 H, H-7), 7.3 1 (s, 1 H, H-4), 3.96 (s, 3 H, OCH₃),2.27 (s, 3 H, 3CH₃).

¹³ C NMR: δ160.20 (s), 140.38 (d), 135.91 (s), 133.21 (d), 133.08 (d),132.73 (s), 131.48(s), 12 7.86(s), 127.14 (d), 116.15 (d), 97.82 (d),58.79 (q), 19.08 (q). Analysis calculated for C₁₅ H₁₄ N₂ O.HCl.0 .25H₂ Orequires: C, 64.5; H, 5.6; N, 10.0%. Found: C, 64.4; H, 5.5; N, 10.2%.

Example 50 5-Hydroxy-6-methyl-1-phenylbenzimidazole Hydrochloride by theMethod of Scheme 3

Demethylation of the ether Example 49 using HBr in acetic acid asdescribed for synthesis of Example 2 gave Example 50 (82%). HCl salt: mp(MeOH/Et₂ O) 239-243° C.

¹ H NMR (D2O): δ9.24 (s, 1 H, H-2), 7.74-7.70 (m, 3 H, Ph), 7.66-7.63(m, 2 H, Ph), 7.49 (s, 1 H, H-7), 7.23 (s, 1 H, H-4), 2.30 (s, 3 H,CH₃).

¹³ C NMR: δ156.80 (s), 140.69 (d), 136.01 (s), 133.25 (d), 133.08 (d),132.55 (s), 130.31 (s), 128.23 (s), 127.38 (d), 116.65 (d), 101.36 (d),18.99 (q). Analysis calculated for C₁₄ H₁₂ N₂ O.HCl requires: C, 64.5;H, 5.0; N, 10.7%. Found: C, 64.2; H, 4.9; N, 10.7%.

Example 51 Methyl 5-methoxy-1-phenylbenzimidazole-6-carboxylateHydrochloride by the Method of Scheme 3

Treatment of 5-methoxy-1-phenylbenzimidazole-6-carboxylic acidhydrochloride with thionyl chloride followed by MeOH, as described forthe synthesis of Example 43, gave Example 51 (91%). HCl salt: mp(MeOH/Et₂ O) 203-205° C.

¹ H NMR (D₂ O): δ9.44 (s, 1 H, H-2), 7.95 (s, 1 H, H-7), 7.73-7.69 (m, 3H, Ph), 7.62-7.59 (m, 2 H, Ph), 7.46 (s, 1 H, H-4), 3.96-3.86 (2s, 6 H,OCH₃).

¹³ C NMR: δ169.72 (s), 160.51 (s), 144.52 (d), 138.12 (s), 135.50 (s),133.44 (d), 133.22 (d), 127.53 (s), 127.02 (d), 121.74 (s), 119.16 (d),100.68 (d), 59.30 (q). Analysis calculated for C₁₆ H₁₄ N₂ O₃ HCl.0.5H₂ Orequires: C, 58.6; H, 4.9; N, 8.6%. Found: C, 58.7; H, 4.5; N, 8.6%.

5-Methoxy-1-phenylbenzimidazole-6-carboxylic acid hydrochloride wasprepared by the method of Scheme 3 as follows: Powdered KMnO₄ (4.08 g,0.026 mol) was added in portions over 12 hours to a refluxing solutionof the free base of 5-methoxy-6-methyl-1-phenylbenzimidazole (Example49) (1.50 g, 6.29 mmol) in 1:1 tert-butanol/water (600 mL), by whichtime no starting material was present. The hot mixture was filteredthrough Celite, washing through with water. The filtrate wasconcentrated under reduced pressure to a volume of conc. 100 mL andwashed with EtOAc. The aqueous portion was carefully neutralized with 3NHCl, chilled to 5° C. for 4 hours, and the precipitated5-methoxy-1-phenylbenzimidazole-6-carboxylic acid (1.08 g, 64%) wasremoved by filtration. HCl salt: mp (MeOH/Et₂ O) 279-281 ° C.

¹ H NMR (D₂ O): 5 9.45 (s, 1 H, H-2), 7.99 (s, 1 H, H-7), 7.71-7.69 (m,3 H, Ph), 7.64-7.61 (m, 2 H, Ph), 7.51 (s, 1 H, H-4), 4.00 (s, 3 H,OCH₃).

¹³ C NMR: δ171.51 (s), 160.37 (s), 144.33 (d), 137.72 (s), 135.53 (s).133.48 (d), 133.21 (d), 127.79 (s), 127.18 (d), 122.83 (s), 119.03 (d),100.57 (d), 59.38 (q). Analysis calculated for C₁₅ H₁₂ N₂ O₃.HCl.0.25H₂O requires: C, 58.3; H, 4.4; N, 9.1%. Found: C, 58.2; H, 4.4; N, 9.0%.

Example 52 5-Hydroxy-1-phenylbenzimidazole-6-carboxylic acidHydrobromide by the Method of Scheme 3

A solution of 5-methoxy-1-phenylbenzimidazole-6-carboxylic acidhydrochloride (0.60 g, 2.24 mmol) in a 33% solution of HBr in glacialacetic acid (50 mL) was refluxed for 30 hours. After cooling to roomtemperature, the resulting precipitate of the hydrobromide salt ofExample 52 was filtered off (0.51 g, 68%): mp 288-290° C. (dec.).

¹ H NMR [(CD₃)₂ SO]: δ9.67 (s, 1 H, H-2), 8.03 (s, 1 H, H-7), 7.82-7.80(m, 2 H, Ph), 7.76-7.71 (m, 2 H, Ph), 7.68-7.64 (m, 1 H, Ph), 7.39 (s, 1H, H-4), 6.00 (br, 2 H, OH).

¹³ C NMR: δ171.15 (s), 158.44 (s), 145.11 (d), 139.64 (s), 133.79 (s),130.29 (d), 129.74 (d), 125.16 (s), 124,70 (d), 114.14 (d), 112.27 (s),102.43 (d). Analysis calculated for C₁₄ H₁₀ N₂ O₃.HBr requires: C, 50.2;H, 3.3; N, 8.4%. Found: C, 49.6; H, 2.9; N, 8.0%.

Example 53 6-Hydroxymethyl-5-methoxy-1-phenylbenzimidazole Hydrochlorideby the Method of Scheme 3

Borane-methyl sulfide complex (1.11 mL of 10.0N, 0.011 mol) was addeddropwise at room temperature under nitrogen to a stirred solution of5-methoxy-1-phenylbenzimidazole-6-carboxylic acid hydrochloride(1.00 g,3.73 mmol) in THF (60 mL). After stirring at this temperature for 3hours, the excess of reagent was destroyed by careful addition of MeOH,followed by water, and the solution was acidified with 3N HCl andstirred at room temperature for a further 30 minutes. The aqueoussolution was washed with EtOAc, then basified with conc. NH₃ solution,extracted with EtOAc, and the organic portion worked up to give an oilysolid which was chromatographed on silica gel. EtOAc eluted foreruns,while MeOH/EtOAc (1:15) eluted the alcohol Example 53 (0.77 g, 81%). HClsalt: mp (MeOH/Et₂ O) 260° C. (dec.).

¹ H NMR (D₂ O): δ9.33 (s, 1 H, H-2), 7.75-7.67 (m, 5 H, Ph), 7.67 (s, 1H, H-7), 7.42 (s, 1 H, H-4), 4.73 (s, 2 H, CH₂ O), 3.99 (s, 3 H, OCH₃).

¹³ C NMR: δ159.85 (s), 141.41 (d), 135.94 (s), 134.25 (s), 133.31 (d),133.14 (d), 132.85 (s), 128.05 (s), 127.34 (d), 115.28 (d), 98.60 (d),62.38 (t), 58.86 (q). Analysis calculated for C₁₅ H₁₄ N₂ O₂.HClrequires: C, 62.0; H, 5.2; N, 9.6%. Found: C, 62.0; H, 5.2; N, 9.5%.

Example 54 5-Methoxy-1-phenylbenzimidazole-6-carboxaldehydeHydrochloride by the Method of Scheme 3

Manganese dioxide (0.20 g, 2.30 mmol) was added in 4 portions over 8hours to a refluxing solution of the free base of the alcohol Example 53(0.10 g, 0.39 mmol) in EtOAc (10 mnL), and refluxing was continued for afurther 10 hours. The mixture was filtered through Celite and thefiltrate percolated through a short column of silica to give thealdehyde (Example 54) (0.098 g, 99%). HCl salt: mp (MeOH//Et₂ O)218-220° C.

¹ H-NMR (D₂ O): δ10.34 (s, 1 H, CHO), 9.57 (s, 1 H, H-2), 8.16 (s, 1 H,H-7), 7.73 (br s, 5 H, Ph), 7.59 (s, 1 H, H-4), 4.08 (s, 3 H, OCH₃).

¹³ C NMR: δ194.97 (d), 163.61 (s), 144.83 (d), 138.88 (s), 135.52 (s),133.67 (d), 133.23 (d), 128.63 (s), 127.55 (d), 127.12 (s), 117.98 (d),100.42 (d), 59.40 (q). Analysis calculated for C₁₅ H₁₄ N₂ O₂.HCI.0.25H₂O requires: C, 61.4; H, 4.6; N, 9.6%. Found: C, 61.4; H, 4.6; N, 9.6%.

Example 55 5-Methoxy-1-(2-thienyl)benzimidazole Hydrochloride by theMethod of Scheme 4

A mixture of 4-methoxy-2-nitroaniline (X: R'=4-OMe) (1.00 g, 5.95 mmol),K₂ CO₃ (1.00 g, 7.23 mmol), CuI (50 mg), and 2-bromothiophene (XI: 2-Br)(5 mL, 0.052 mol) was refluxed under nitrogen with stirring for 18hours. Excess bromothiophene was removed under reduced pressure, and theresidue was partitioned between EtOAc and water and filtered throughCelite. The organic portion was worked up to give an oil which waschromatographed on silica gel. EtOAc/petroleum ether (1:9) eluted4-methoxy-2-nitro-N-(2-thienyl)aniline (XII: R=4-OMe, 2-thienyl) (0.37g, 25%): mp (aqueous EtOH) 108-110° C.

¹ H NMR (CDCl₃): δ9.17 (br, 1 H, NH), 7.37 (dd, J=1.8, 1.6 Hz, 1 H),7.15 (dd, J=5.7, 1.4 Hz, 1 H), 7.09 (d, J=1.4 Hz, 1 H), 7.08 (s, 1 H),6.98 (dd, J=5.7, 3.6 Hz, 1 H), 6.88 (m, 1 H), 3.82 (s, 3 H, OCH₃).

¹³ C NMR: δ151.39 (s), 141.70 (s), 139.57 (s), 126.41 (d), 126.20 (d),123.47 (d), 122.80 (d), 118.21 (s), 117.65 (d), 106.78 (d), 55.59 (q).Analysis calculated for C₁₁ H₁₀ N₂ O₃ S requires: C, 52.8; H, 4.0; N,11.2%. Found: C, 53.1;H,4.1; N, 11.3%.

Hydrogenation of the above thienylaniline over 5% Pd-C, followed byreaction with formamidine acetate, gave Example 55 (82%) as thehydrochloride salt: mp (MeOH/Et₂ O) 169-172° C.

¹ H NMR (D₂ O): δ9.36 (s, 1 H, H-2), 7.68 (dd, J=1.4, 6.8 Hz, 1 H,H-5'), 7.65 (d, J=9.2 Hz, 1 H, H-7), 7.49 (dd, J=3.9, 1.4 Hz, 1 H,H-4'), 7.37 (d, J=2.3 Hz, 1 H, H-4), 7.27 (dd, J=6.8, 3.9 Hz, 1 H,H-3'), 7.25 (dd, J=9.2, 2.3 Hz, 1 H, H-6), 3.94 (s, 3 H, OCH₃).

¹³ C NMR: δ161.58(s), 143.20(d), 135.10(s), 134.57(s), 129.68(s),129.58(d), 129.47 (d), 128.24 (d), 119.94 (d), 116.50 (d), 100.07 (d),58.87 (q). Analysis calculated for C₁₂ H₁₀ N₂ OS.HCl.0.5H₂ O requires:C, 52.3; H, 4.4; N, 10.2%. Found: C, 52.2; H, 4.3; N, 10.2%.

Example 56 5-Methoxy-1-(3-thienyl)benzimidazole Hydrochloride by theMethod of Scheme 4

Reaction of 4-methoxy-2-nitroaniline (X: R=4-OMe) and 3-bromothiophene(XI: R'=3-Br) as described above for Example 55, but for only 6 hours,gave 4-methoxy-2-nitro-N-(3-thienyl)aniline (XIV: R'=4-OMe, 3-thienyl)(71%): mp (EtOAc/petroleum ether) 121-123° C.

1H NMR (CDCl₃): δ9.31 (br, 1 H, NH), 7.63 (d, J=3.0 Hz, 1 H, H-3), 7.36(dd, J=5.1, 3.2 Hz, 1 H, H-4'), 7.18 (d, J=9.4 Hz, 1 H, H-6), 7.09 (dd,J=9.4, 3.0 Hz, 1 H, H-5), 7.05 (dd, J=3.2, 1.4 Hz, 1 H, H-2'), 7.01 (dd,J=5.1, 1.4 Hz, 1 H, H-5'), 3.82 (s, 3 H, OCH₃).

¹³ C NMR: δ151.01 (s), 138.79 (s), 138.71 (s), 132.12 (s), 126.55 (d),126.06 (d), 124.74 (d), 117.63 (d), 115.05 (d), 106.82 (d), 55.86 (q).Analysis calculated for C₁₁ H₁₀ N₂ O₃ S: C, 52.8; H, 4.0; N, 11.2%.Found: C, 52.8; H, 4.2; N, 11.1%.

Hydrogenation of the above thienylaniline over 5% Pd-C followed byreaction with formamidine acetate gave Example 56 (77%) as thehydrochloride salt: mp (MeOH/Et₂ O) 219-221° C. (dec.).

¹ H NMR (D₂ O): δ9.36 (s, 1 H, H-2), 7.88 (dd, J=3.1, 1.5 Hz, 1 H,H-2'), 7.81 (dd, J=5.1, 3.1 Hz, 1 H, H-4'), 7.62 (d, J=9.2 Hz, 1 H,H-7), 7.44 (dd, J=5.1, 1.5 Hz, 1 H, H-5'), 7.32 (d, J=2.3 Hz, 1 H, H-4),7.20 (dd, J=9.2, 2.3 Hz, 1 H, H-6), 3.93 (s, 3 H, OCH₃).

¹³ C NMR: δ161.43 (s), 141.67 (d), 134.40 (s), 133.32 (s), 131.49 (d),128.34 (s), 125.40 (d), 123.46 (d), 119.77 (d), 116.65 (d), 99.76 (d),58.84 (q). Analysis calculated for C₁₂ H₁₀ N₂ SO.HCl requires: C, 54.0;H, 4.2; N, 10.5%. Found: C, 54.1; H, 4.2; N, 10.6%.

Example 57 5-(4-Hydroxybutoxy)-1-phenylbenzimidazole Hydrochloride bythe Method of Scheme 3

A mixture of 5-hydroxy-1-phenylbenzimidazole (0.31 g, 1.47 mmol), CsCO₃(0.58 g, 1.77 mmol), and 4-chlorobutanol (0.16 mL, 1.62 mmol) in DMF (10mL) was warmed at 115° C. for 18 hours, then concentrated to drynessunder reduced pressure. The residue was partitioned between EtOAc andwater, then filtered, and the organic portion was worked up andchromatographed on silica gel. EtOAc eluted a trace of startingmaterial, while MeOH/EtOAc (1:9) gave the alcohol (Example 57) (0.34 g,82%). HCl salt: mp (MeOH/EtOAc) 205-207° C.

¹ H NMR (D₂ O): δ9.37 (s, 1 H, H-2), 7.76-7.67 (m, 5 H, Ph), 7.66 (d,J=9.2 Hz, 1 H, H-7), 7.41 (d, J=1.6 Hz, 1 H, H-4), 7.28 (dd, J=9.2, 1.6Hz, 1 H, H-6), 4.19 (t, J=6.4 Hz, 2 H, CH₂ O), 3.69 (t, J=6.5 Hz, 2 H,CH₂ O), 1.93-1.86 (m, 2 H, CH₂), 1.79-1.71 (m, 2 H, CH₂). ¹³ C NMRδ160.78 (s), 141.83 (d), 135.95 (s), 134.60 (s), 133.34 (d), 133.13 (d),128.82 (s), 127.44 (d), 120.19 (d), 116.79 (d), 100.85 (d), 71.89 (t),64.11 (t), 30.67 (t), 27.62 (t). Analysis calculated for C₁₇ H₁₈ N₂O₂.HCl.0.5H₂ O requires: C, 62.3; H, 6.2; N, 8.6%. Found: C, 61.8; H,5.8; N, 8.4%.

Example 58 5-[4-(N,N-Dimethylamino)butoxy]-1-phenylbenzimidazoleDihydrochloride by the Method of Scheme 3

Methanesulfonyl chloride (0.27 mL, 3.31 mmol) was added dropwise at 20°C. to a stirred solution of 5-(4-hydroxybutoxy)-1-phenylbenzimidazolehydrochloride (0.85 g, 3.01 mmol) and Et₃ N (0.50 mL, 3.61 mmol) in CH₂Cl₂ (50 mL). After 5 minutes the resulting solution was evaporated todryness under reduced pressure to give the crude mesylate. Half of thiswas dissolved in MeOH (20 mL) and dimethylamine (5 mL of a 40% aqueoussolution), and the mixture was warmed in a pressure vessel at 80° C. for15 hours. After concentration to dryness the residue was partitionedbetween EtOAc and water, and the organic portion was chromatographed onalumina. EtOAc eluted foreruns while MeOH/EtOAc (1:9) gave thedimethylamine (Example 58) (0.32 g, 65%). DiHCl salt: mp (MeOH/Et₂ O)88° C. (hygroscopic).

¹ H NMR (D₂ O): δ9.31 (s, 1 H, H-2), 7.75-7.70 (m, 5 H, Ph), 7.70 (d,J=9.2 Hz, 1 H, H-7), 7.43 (d, J=2.3 Hz, 1 H, H-4), 7.30 (dd, J=9.2, 2.3Hz, 1 H, H-6), 4.23 (t, J=5.7 Hz, 2 H, CH₂ O),3.25 (t, J=7.0 Hz, 2 H,CH₂ N⁺), 2.91 (s, 6 H, ⁺ NMe₂), 2.00-1.91 (m, 4 H, CH₂). HRMS (EI): C₁₉H₂₃ N₃ O requires: M⁺ 309.1841. Found: M⁺ 309.1810

Example 59 5-[4-(4-Morpholino)butoxy]-1]phenylbenzimidazoleDihydrochloride by the Method of Scheme 3

The crude mesylate prepared in Example 58 (1.5 mmol) was dissolved inmorpholine (10 mL) and refluxed for 30 minutes. After removal of excessmorpholine under reduced pressure, the residue was partitioned betweenbrine and EtOAc and the organic portion was worked up andchromatographed on alumina. EtOAc eluted the morpholide (Example 59)(0.27 g, 49%). DiHCl salt: mp (MeOH/Et₂ O) 100-103° C. (hygroscopicpowder).

¹ H NMR (D₂ O): δ9.40 (s, 1 H, H-2), 7.75-7.72 (m, 5 H, Ph), 7.70 (d,J=9.2 Hz, 1 H, H-7), 7.45 (d, J=2.3 Hz, 1 H, H-4), 7.32 (dd, J=9.2, 2.3Hz, 1 H, H-6), 4.24 (t, J=5.9 Hz, 2 H, CH₂ O), 4.17 (dd, J=13.2, 3.4Hz,2 H, CH₂ O), 3.90-3.82 (m, 2 H, CH₂ O), 3.61-3.59 (m, 2 H, CH₂ N⁺), 3.32(t, J=6.9 Hz, 2 H, CH₂ N⁺), 3.28 3.21 (m, 2 H, CH₂ N⁺).

¹³ C NMR δ160.69 (s), 141.91 (d), 135.95 (s), 134.54 (s), 133.37 (d),133.14 (d), 128.88 (s), 127.50 (d), 120.17 (d), 116.89 (d), 100.80 (d),71.02 (t), 66.54 (t), 59.64 (t), 54.39 (t), 28.13 (t), 22.89 (t). HRMS(CI). C21H₂₆ N₃ O₂ requires: [M+H] 352.2025. Found [M+H] 352.2103.

Example 60 4,5-Dimethoxy-1-phenylbenzimidazole Hydrochloride by theMethod of Scheme 1

A solution of 3,4-dimethoxy-2-nitrobenzoic acid (6.26 g, 0.027 mol),SOCl₂ (10.0 mL) and DMF (1 drop) in 1,2-dichloroethane (100 mL) wasrefluxed for 2 hours, then concentrated to dryness under reducedpressure. The resulting crude acid chloride was dissolved in acetone (50mL), cooled to 5° C. and treated in one portion with a solution ofexcess sodium azide (10.0 g) in water (20 mL). After vigorous stirringfor 10 minutes the mixture was poured into water (300 mL) and theresultant precipitate of acyl azide was filtered off and washed wellwith water. The crude solid was slurried in acetic acid (300 mL) andwater (30 mL) and heated slowly to reflux. After refluxing for 2 hoursthe solvents were removed under reduced pressure and the residue wasslurried in hot EtOH and filtered. The filtrate was concentrated todryness and the residue was triturated in EtOAc and worked up to give3,4-dimethoxy-2-nitroaniline (12.96 g, 55%): mp (aqueous EtOH) 65-67° C.

¹ H NMR (CDCl₃) δ6.94 (d, J=9.1 Hz, 1 H, H-5), 6.49 (d, J=9.1 Hz, 1 H,H-6), 4.44 (br, 2 H, NH₂) 3.96, 3.82 (2 s, 6 H, OCH₃).

¹³ C NMR δ145.03 (s), 144.00 (s), 139.45 (s), 135.73 (s), 119.15 (d),111.75 (d), 61.94 (OCH₃), 57.33 (OCH₃). Analysis calculated for C₈ H₁₀N₂ O₄ requires: C, 48.5; H, 5.1; N, 14.1%. Found: C, 48.8; H, 5.1; N,13.8%.

A mixture of the above dimethoxynitroaniline (1.40 g, 7.06 mmol), K₂ CO₃(1.50 g, 0.011 mol), Cul (0.10 g) and bromobenzene (15 mL) was refluxedwith vigorous stirring for 8 hours. After removal of excess bromobenzeneunder reduced pressure the residue was partitioned between EtOAc andwater, and the organic portion was worked up to give an oil which waschromatographed on silica gel. Petroleum ether eluted a littlebromobenzene while EtOAc/petroleum ether (1:9) gave3,4-dimethoxy-2-nitrodiphenylamine (0.46 g, 24%): mp (EtOAc/petroleumether) 68-69° C.

¹ H NMR (CDCl₃) δ7.28 (m, 2 H, Ar), 7.06-6.95 (m, 5 H, Ar), 6.68 (br s,1 H, NH), 4.00, 3.87 (2 s, 6 H, OCH₃).

¹³ C NMR δ146.99 (s), 143.30 (s), 141.61 (s), 137.40 (s), 131.47 (s),129.48 (d), 122.36 (d), 119.19 (d), 117.12 (d), 113.70 (d), 62.09(OCH₃), 56.92 (OCH₃). Analysis calculated for C₁₄ H₁₄ N₂ O₄ requires: C,61.1; H, 5.1, N, 10.2%. Found: C, 61.1; H, 5.1; N, 10.0%.

Reduction of the above diphenylamine with H₂ /Pd--C followed by reactionof the resulting diamino compound with formamidine acetate, gave thediether (Example 60) (76%). HCl salt: mp (MeOH/Et₂ O) 180-183° C. (dec).

¹ H NMR (D₂ O) δ9.37 (s, 1 H, H-2), 7.72 (m, 3 H, Ph), 7.64 (m, 2 H,Ph), 7.41 (d, J=9.1 Hz, 1 H, H-7), 7.37 (d, J=9.1 Hz, 1 H, H-6), 4.08,4.00 (2s, 6 H, OCH₃).

¹³ C NMR δ152.62 (s), 142.77 (d), 137.88 (s), 135.85 (s), 133.26 (d),133.13 (d), 129.82 (s), 128.86 (s), 127.09 (d), 117.20 (d), 111.09 (d),64.50 (OCH₃), 59.94 (OCH₃). Analysis calculated for C₁₅ H₁₄ N₂O₂.HCl.0.5H₂ O requires: C, 60.1; H, 5.4; N, 9.4%. Found: C, 60.3; H,5.5; N, 9.4%.

Example 61 4,5-Dihydroxy-1-phenylbenzimidazole Hydrochloride by theMethod of Scheme 3

A mixture of the free base of the diether (Example 60) (60 mg, 0.23mmol) and lithium methanethiolate (0.20 g, 3.77 mmol) in dry DMF (10 mL)was warmed at 120° C. under an atmosphere of nitrogen for 4 hours. Thecooled solution was carefully neutralised with conc. HCl then pouredinto brine and extracted into EtOAc. The extract was worked up andchromatographed on silica gel. EtOAc gave foreruns while MeOH/EtOAc(1:19) eluted the diphenol (Example 61) (46 mg, 88%). HCl salt: mp(MeOH/Et₂ O) 241-244° C.

¹ H NMR (D₂ O) δ9.27 (s, 1 H, H-2), 7.72-7.59 (m, 5 H, Ph), 7.18 (d,J=9.0 Hz, 1 H, H-7), 7.11 (d, J=9.0Hz, 1 H, H-6).

¹³ C NMR δ144.99 (s), 141.76 (d), 136.08 (s), 134.41 (s), 133.16 (d),133.07 (d), 129.60 (s), 127.28 (d), 125.89 (s), 120.01 (d), 106.96 (d).Analysis calculated for C₁₃ H₁₀ N₂ O₂.HCl requires: C, 59.4; H, 4.2; N,10.7%. Found: C, 59.3; H, 4.2; N, 10.7%.

Example 62 5-Hydroxy-4-methoxy-1-phenylbenzimidazole Hydrochloride bythe Method of Scheme 3

Boron tribromide (1.12 mL of a 1N solution in CH₂ Cl₂, 1.12 mmol) wasadded under nitrogen at 5° C. to a solution of the diether (Example 60)(0.24 g, 1.12 mmol) in CH₂ Cl₂ (20 mL). After 1 hour at thistemperature, followed by 2 hours at room temperature, 2N NaOH (10 mL)was added and the mixture was stirred for 1 hour. The aqueous layer wascarefully neutralised with 2N HCl and extracted with EtOAc, and theextract was worked up and chromatographed on silica gel. Elution withEtOAc/petroleum ether (1:1) gave Example 62 (26 mg, 12%). HCl salt: mp(MeOH//t₂ O) 220-222° C.

¹ H NMR (CDCl₃) δ8.00 (s, 1 H, H-2), 7.58-7.43 (m, 5 H, Ph), 7.08 (d,J=8.7 Hz, 1 H, H-7), 6.99 (d, J=8.7 Hz, 1 H, H-6), 5.66 (br, 1 H, OH),4.44 (s, 3 H, OCH₃).

¹³ C NMR δ142.54 (s), 141.45 (d), 136.77 (s), 136.57 (s), 136.29 (s),135.51 (s), 130.02 (d), 128.03 (d), 124.03 (d), 112.43 (d), 103.99 (d),61.10 (OCH₃). Analysis calculated for C₁₄ H₁₂ N₂ O₂.HCl requires: C,60.8; H, 4.7; N, 10.1%. Found: C, 61.3; H, 4.5; N, 9.5%.

Example 63 4-Hydroxy-5-methoxy-1-phenylbenzimidazole Hydrochloride bythe Method of Scheme 3

Further elution with of the column chromatography in Example 62 withEtOAc gave Example 63 (0.11 g, 50%): mp (EtOAc/petroleum ether) 196-197°C.

¹ H NMR (CDCl₃) δ8.16 (s, 1 H, H-2), 7.60-7.45 (m, 5 H, Ph), 7.06 (d,J=8.7 Hz, 1 H, H-7), 6.99 (d, J=8.7 Hz, 1 H, H-6), 3.98 (s, 3 H, OCH₃),1.27 (br, 1 H, OH).

Analysis calculated for C₁₄ H₁₂ N₂ O₂ requires: C, 70.0; H, 5.0; N,11.7%. Found: C, 70.3; H, 5.2; N, 11.7%.

Example 64 4-Bromo-5-Hydroxy-1-phenylbenzimidazole Hydrochloride by theMethod of Scheme 3

A solution of N-bromosuccinimide (0.72 g, 4.04 mmol) in DMF (5 mL) wasadded dropwise at room temperature to a solution of5-hydroxy-1-phenylbenzimidazole (0.85 g, 4.04 mmol) in DMF (30 mL).After 1 hour the solution was diluted with aqueous NaCl and extractedinto EtOAc, and the extract was worked up to give a solid which waschromatographed on silica gel. Elution with EtOAc/petroleum ether (1:1)gave foreruns, while EtOAc eluted Example 64 (0.88 g, 75%). HCl salt: mp(MeOH/Et₂ O) 244-246° C.

¹ H NMR δ(free base in CDCl₃) 8.12 (s, 1 H, H-2), 7.60-7.56 (m, 2 H,Ph), 7.50-7.46 (m, 3 H, Ph), 7.36 (d, J=Hz, 1 H, H-7), 7.08 (d, J=8.8Hz, 1 H, H-6), 5.85 (br, 1 H, OH).

¹³ C NMR δ149.30 (s), 142.88 (d), 135.91 (s), 130.14 (d), 128.44 (s),128.41 (d), 124.11 (s), 124.03 (d), 113.18 (d), 110.33 (d), 99.66(s).

Analysis calculated for C₁₃ H₁₉ BrN₂ O.HCl requires: C, 48.0; H, 3.1; N,8.6%.

Found: C, 47.8; H, 3.3; N, 8.6%.

Example 65 4-Bromo-5-allyloxy-1-phenylbenzimidazole by the Method ofScheme 3

A solution of the above 5-phenol (Example 64) (0.92 g, 3.18 mmol) in DMF(4 mL) was added under nitrogen to s stirred suspension of NaH (0.15 gof a 60% dispersion in oil, 3.81 mmol). After 5 minutes, allyl bromide(0.30 mL, 3.50 mmol) was added and the solution was stirred at roomtemperature for 1 hour and at 50° C. for 30 minutes. After dilution withbrine the mixture was extracted with diethyl ether. The ether portionwas washed with 2N NaOH and worked up to give Example 65 (1.03 g, 99%).Mp: (EtOAc/petroleum ether) 95-96° C.

¹ H NMR (CDCl₃) δ8.15 (s, 1 H, H-2), 7.60-7.56 (m, 2 H, Ph), 7.50-7.46(m, 3 H, Ph), 7.38 (d, J=8.8 Hz, 1 H, H-7), 7.02 (d, J=8.8 Hz, 1 H,H-6), 6.16-6.09 (m, 1 H, ═CH), 5.16-5.47 (m, 1 H, ═CHH), 5.32-5.29 (m, 1H, ═CHH), 4.69-4.67 (m, 2 H, OCH₂).

¹³ C NMR δ151.86 (s), 143.94 (s), 143.23 (d), 135.99 (s), 133.16 (d),130.13 (d), 129.35 (s), 128.34 (d), 124.00 (d), 117.81 (t), 112.91 (d),109.39 (d), 103.77 (s), 71.82 (t).

Analysis calculated for C₁₆ H₁₃ BrN₂ O requires: C, 58.4; H, 4.0; N,8.5%.

Found: C, 58.4; H, 4.0; N, 8.7%.

Example 66 5-(Methylthio)-1-phenylbenzimidazole Hydrochloride by theMethod of Scheme 3

A solution of 5-hydroxy-1-phenylbenzimidazole (Example 14) (0.88 g, 4.10mmol) in 1:1 THF/DMF (15 mL) was added dropwise under nitrogen to astirred suspension of NaH (0.24 g of a 50% dispersion in oil, 5.00 mmol)in THF (10 mL). After 10 minutes a solution of dimethylthiocarbamylchloride (0.57 g, 4.6 mmol) in THF (5 mL) was added and the solution waswarmed at 60° C. for 1 hour. The cooled solution was partitioned betweenether and 2N KOH and the organic portion was worked up andchromatographed on silica. Elution with EtOAc/petroleum ether (7:3) gavethe O-benzimidazolyl dimethylthiocarbamate (0.76 g, 65%), mp(EtOAc/petroleum ether) 173-174.5° C.

¹ H NMR [(CD₃)₂ SO]: d 8.61 (s, 1H, H-2), 7.72-7.62 (m, 4H, Ph), 7.60(d, J=8.7 Hz, 1H, H-7), 7.54-7.49 (m, 1H, Ph), 7.45 (d, J=2.2 Hz, 1H,H-4), 7.04 (dd, J=8.7, 2.2 Hz, 1H, H-6), 3.39, 3.36 (2s, each 3H, NMe₂).

¹³ C NMR: 187.03 (s), 149.51 (s), 144.27 (d), 143.78 (s), 135.74 (s),130.78 (s), 130.03 (d), 127.79 (d), 123.60 (d), 119.02 (d), 113.55 (d),110.32 (d), 42.80 (q), 38.41 (q).

A solution of this thiocarbamate (0.40 g, 1.40 mmol) in dry sulfolane(20 mL) was refluxed under nitrogen for 5 hours. The cooled solution waspoured into brine, extracted with EtOAc and worked up to give an oilwhich was chromatographed on silica. EtOAc/petroleum ether (1:1) elutedresidual sulfolane while EtOAc gave the S-benzimidazolyldimethylthiocarbamate (0.36 g, 90%) as an oil which was hydrolyseddirectly. A solution of this crude thiocarbamate (0.36 g, 1.26 mmol) inMeOH (50 mL) and 6 N KOH (5 mL) was refluxed for 6 hours. After removalof the MeOH in vacuo the residue was partitioned between ether andwater. The aqueous portion was carefully neutralised with conc. HCl,extracted with EtOAc and the extract worked up to give an oil which waschromatographed on silica. EtOAc eluted 5-mercapto-1-phenylbenzimidazoleas an unstable oil (0.16 g, 56%). HCl salt: mp (MeOH/Et₂ O) 180-184° C.

¹ NMR (D₂ O): d 9.44 (s, 1H, H-2), 7.84 (d, J=2.0 Hz, 1H, H-4),7.75-7.65 (m, 5H, Ph), 7.63 (d, J=8.7 Hz, 1H, H-7), 7.54 (dd, J=8.7, 2.0Hz, 1H, H-6).

¹³ C NMR: 142.48 (d), 135.70 (s), 134.18 (s), 133.94 (s), 133.48 (d),133.16 (d), 133.12 (s), 130.86 (d), 127.52 (d), 116.70 (d), 116.42 (d).

A solution of the mercaptobenzimidazole (0.14 g, 0.61 mmol) in 1:1THF/DMF (5 mL) was added dropwise under nitrogen to a stirred suspensionof NaH (35 mg of a 50% dispersion in oil, 0.73 mmol). After 5 minutesmethyl iodide (42 mL, 0.67 mmol) was added and the solution was stirredat 20° C. for 3 hours, then poured into 1 N NaOH and extracted withether. The extract was chromatographed on silica, eluting withEtOAc/petroleum ether (1:1) to give Example 66 as an oil (0.09 g, 61%).HCl salt: mp (MeOH/Et₂ O) 192-195° C.

¹ NMR (D₂ O): d 9.45 (s, 1H, H-2), 7.76-7.72 (m, 3H, Ph), 7.67-7.63 (m,3H, Ph and H-4), 7.57 (d, J=8.9 Hz, 1H, H-7), 7.43 (dd, J=8.9, 1.6 Hz,1H, H-6), 2.59 (s, 3H, SCH₃).

¹³ C NMR: 142.09 (d), 141.36 (s), 135.63 (s), 134.18 (s), 133.41 (d),133.17 (d), 131.55(s), 128.46(d), 127.16(d), 115.91 (d), 113.36(d),17.57(q).

Purification of Tyrosine Kinases

Epidermal Growth Factor Receptor (EGFr)

Human EGF receptor tyrosine kinase is isolated from A431 humanepidermoid carcinoma cells by the following method. Cells were grown inroller bottles in 50% Dulbecco's Modified Eagle medium and 50% HAM F-12nutrient media (Gibco) containing 10% fetal calf serum. Approximately10⁹ cells are lysed in two volumes of buffer containing 20 mM2-(4N-[2-hydroxymethyl]piperazin-1-yl)ethanesulfonic acid (Hepes), pH7.4, 5 mM ethylene glycol bis(2-aminoethyl ether) N,N,N',N'-tetraaceticacid (EGTA), 1% Triton X-100, 10% glycerol, 0.1 mM sodium orthovanadate,5 mM sodium fluoride, 4 mM pyrophosphate, 4 mM benzamide, 1 mMdithiothreitol (DTT), 80 μg/mL aprotinin, 40 μg/mL leupeptin, and 1 mMphenylmethylsulfonyl fluoride (PMSF). After centrifugation at 25,000 ×gfor 10 minutes, the supernatant is applied to a fast Q sepharose column(Pharmacia) and eluted with a linear gradient from 0.1 M NaCl to 0.4 MNaCl in 50 mM Hepes, 10% glycerol, pH 7.4. Enzyme active fractions arepooled, divided into aliquots and stored at -100° C.

Platelet Derived Growth Factor Receptor (PDGFr) and Fibroblast GrowthFactor Receptor (FGFr)

Full length cDNAs for the mouse PDGF-β and human FGF-1 (flg) receptortyrosine kinases were obtained from J. Escobedo and are prepared asdescribed in J. Biol. Chem., 262: 1482-1487 (1991). PCR primers aredesigned to amplify a fragment of DNA that codes for the intracellulartyrosine kinase domain. The fragment is melded into a baculovirusvector, cotransfected with AcMNPV DNA, and the recombinant virusisolated. SF9 insect cells are infected with the virus to overexpressthe protein, and the cell lysate is used for the assay.

Other Kinases

c-Src kinase is purified from baculovirus infected insect cell lysatesusing an antipeptide monoclonal antibody directed against the N-terminal2-17 amino acids as described previously by Fry, et al, Anticancer DrugDesign. 9: 331-351 (1994). Protein kinase C (PKC) is obtained as a ratbrain preparation from Promega.

Kinase Assays

EGFr

Enzyme assays for IC₅₀ determinations are performed in 96-well filterplates (Millipore MADVN6550). The total volume is 0.1 mL containing 20mM Hepes, pH 7.4, 50 μM sodium vanadate, 40 mM magnesium chloride, 10 μMATP containing 0.5 μCi of [³² P]ATP, 20 μg of polyglutamic acid/tyrosine(Sigma Chemical Co, St. Louis, Mo.), 10 ng of EGF receptor tyrosinekinase and appropriate dilutions of inhibitor. All components except theATP are added to the well and the plate is incubated with shaking for 10minutes at 25° C. The reaction is started by adding [³² P]ATP and theplate is incubated at 25° C. for 10 minutes. The reaction is terminatedby addition of 0.1 mL of 20% trichloroacetic acid (TCA). The plate iskept at 4° C. for at least 15 minutes to allow the substrate toprecipitate. The wells are then washed 5 times with 0.2 mL of 10% TCA,and ³² p incorporation is determined with a Wallac beta plate counter.

PDGFr and FGFr

The assay is performed in 96-well plates (100 μL/incubation/well), andconditions are optimized to measure the incorporation of ³² p from [γ³²P]-ATP into a glutamate-tyrosine copolymer substrate. Briefly, to eachwell is added 82.5 μL of incubation buffer containing 25 mM Hepes (pH7.0), 150 mM NaCl, 0.1% Triton X-100, 0.2 mM PMSF, 0.2 mM sodiumvanadate, 10 mM manganese chloride, and 750 μg/mL of poly (4:1)glutamate-tyrosine followed by 2.5 μL of inhibitor and 5 μL of enzymelysate (7.5 μg/μL FGFR-TK or 6.0 μg/μL PDGFR-TK) to initiate thereaction. Following a 10 minute incubation at 25° C., 10 μL of [γ³²P]-ATP (0.4 μCi plus 50 μM ATP) is added to each well and samples areincubated for an additional 10 minutes at 25° C. The reaction isterminated by the addition of 100 μL of 30% trichloroacetic acid (TCA)containing 20 mM sodium pyrophosphate and precipitation of material ontoglass fiber filter mats (Wallac). Filters are washed 3 times with 15%TCA containing 100 mM sodium pyrophosphate and the radioactivityretained on the filters counted in a Wallac 1250 Betaplate reader.Nonspecific activity is defined as radioactivity retained on the filtersfollowing incubation of samples with buffer alone (no enzyme). Specificenzymatic activity is defined as total activity (enzyme plus buffer)minus nonspecific activity. The concentration of a compound thatinhibited specific activity by 50% (IC₅₀) is determined based on theinhibition curve.

c-Src

The antibody, covalently linked to 0.65-μm latex beads, is added to asuspension of insect cell lysis buffer comprised of 150 mM NaCl, 50 mMTris pH 7.5, 1 mM DTT, 1% NP-40, 2 mM EGTA, 1 mM sodium vanadate, 1 mMPMSF, 1 μg/mL each of leupeptin, pepstatin, and aprotinin. Insect celllysate containing the c-Src protein is incubated with these beads for 3to 4 hours at 4° C. with rotation. At the end of the lysate incubation,the beads are rinsed 3 times in lysis buffer, resuspended in lysisbuffer containing 10% glycerol, and frozen. These latex beads arethawed, rinsed 3 times in assay buffer which is comprised of 40 mM trispH 7.5, 5 mM magnesium chloride, and suspended in the same buffer. In aMillipore 96-well plate with a 0.65 μm polyvinylidine membrane bottomare added the reaction components: 10-μL c-Src beads, 10 μL of 2.5 mg/mLpoly glutamate-tyrosine substrate, 5 μM ATP containing 0.2 μCi labeled³² P-ATP, 5 μL DMSO containing inhibitors or as a solvent control, andbuffer to make the final volume 125 μL. The reaction is started at roomtemperature by addition of the ATP and quenched 10 minutes later by theaddition of 125 μL of 30% TCA, 0.1 M sodium pyrophosphate for 15 minuteson ice. The plate is then filtered and the wells washed with two 250-μLaliquots of 15% TCA, 0.1 M pyrophosphate. The filters are punched,counted in a liquid scintillation counter, and the data examined forinhibitory activity in comparison to a known inhibitor such aserbstatin. The method is described more fully in J. Med. Chem., 37:598-609 (1994).

Cascade Assay for Inhibitors of the MAP Kinase Pathway (APK Assay)

Incorporation of ³² P into myelin basic protein (MBP) is assayed in thepresence of a glutathione S-transferase fusion protein containing p44MAPkinase (GST-MAPK) and a glutathione S-transferase fusion proteincontaining p45MEK (GST-MEK). The assay solution contains 20 mM HEPES, pH7.4, 10 mM magnesium chloride, 1 mM manganese chloride, 1 mM EGTA, 50 μM[γ³² P]ATP, 10 μg GST-MEK, 0.5 μg GST-MAPK and 40 μg MBP in a finalvolume of 100 μL. Reactions are stopped after 20 minutes by addition oftrichloroacetic acid and filtered through a GF/C filter mat. ³² Pretained on the filter mat is determined using a 1205 betaplate.Compounds are assessed at 10 μM for ability to inhibit incorporation of³² P.

To ascertain whether compounds are inhibiting GST-MEK or GST MAPK, twoadditional protocols are employed. In the first protocol, compounds areadded to tubes containing GST MEK, followed by addition of GST-MAPK, MBPand [γ³² P]ATP. In the second protocol, compounds are added to tubescontaining both GST-MEK and GST-MAPK, followed by MBP and [γ³² P]ATP.Compounds that show activity in both protocols are scored as MAPKinhibitors, while compounds showing activity in only the first protocolare scored as MEK inhibitors.

Other Kinases

An assay using the intracellular kinase domains of insulin receptor(INSr) is performed as described for the EGF receptor except that 10 mMmanganese chloride is included in the reaction. The PKC assay isperformed as previously described by Martiny-Baron, et al., J. Biol.Chem. 268: 9194-9197 (1993).

Table 1 provides inhibition data against receptor kinases for compoundsof the present invention.

                  TABLE 1                                                         ______________________________________                                                PDGF    FGF     c-Src EGFr  INSr APK  PKC                             Example IC.sub.50                                                                             IC.sub.50                                                                             IC.sub.50                                                                           IC.sub.50                                                                           IC.sub.50                                                                          IC.sub.50                                                                          IC.sub.50                       Number  (μM) (μM) (μM)                                                                             (μM)                                                                             (μM)                                                                            (μM)                                                                            (μM)                         ______________________________________                                         1      9.3     >50     >50   >50   >50  ND   ND                               2      3.8     50      >50   ND    >50  ND   ND                               3      3.6     >50     >50   >50   >50  ND   ND                               4      6.8     >50     >50   ND    >50  ND   ND                               5      12      >50     >50   ND    >50  ND   ND                               6      1.8     5.8     >50   ND    >50  ND   ND                               7      7.2     >50     >50   >50   >50  ND   ND                               8      5.6     >50     ND    ND    ND   ND   ND                               9      13.5    >50     >50   ND    ND   ND   ND                              10      12.3    50      >50   ND    >50  ND   ND                              11      10.6    >50     >50   >50   >50  ND   ND                              12      4.4     >50     >50   50    >50  ND   ND                              13      0.4     22      >50   <50   >50   5   >50                             14      0.4     6.4     >50   >50   >50  ND   ND                              15      4.0     51      ND    ND    ND   ND   ND                              16      9.3     28      50    >50   >50  ND   ND                              17      0.8     6.6     >50   >50   >50  ND   ND                              18      1.6     9.9     <50   >50   >50  ND   ND                              19      2.7     36      >50   >50   >50  ND   ND                              20      0.9     14      ND    ND    ND   ND   ND                              21      8.4     50      50    <50   >50  ND   ND                              22      10      49      >50   ND    >50  ND   ND                              23      3.3     32      <50   ND    >50  ND   ND                              24      2.4     41      >50   ND    >50  ND   ND                              25      5.3     37      >50   ND    ND   ND   ND                              26      11.7    >50     >50   <50   >50  ND   ND                              27      1.8     26      >50   >50   >50  ND   ND                              28      0.2     26      >50   ND    >50  ND   ND                              29      0.3     35      >50   ND    >50  ND   ND                              30      <50     50      >50   ND    >50  ND   ND                              31      1.4     >50     >50   ND    >50  ND   ND                              32      0.6     35      >50   ND    >50  ND   ND                              33      0.3     15      >50   ND    ND   ND   ND                              34      0.3     18      50    ND    ND   ND   ND                              35      0.7     27      <50   ND    >50  ND   ND                              36      1.5     45      29    ND    >50  ND   ND                              37      0.2     4.1     14    >40   >50  27   >50                             38      0.7     28      >50   >40   >50  45   >50                             39      0.2     9.8     >50   >40   >50  18   >50                             40      6.4     >50     ND    ND    ND   ND   ND                              41      2.1     >50     >50   >50   <50  ND   ND                              42      5.4     >50     ND    ND    ND   ND   ND                              43      13      >50     >50   >50   >50  ND   ND                              44      9.6     >50     ND    ND    ND   ND   ND                              45      0.3     18      >50   <40   >50  37   >50                             46      1.2     25      40    ND    >50  ND   ND                              47      2.3     11      >50   ND    >50  ND   ND                              48      2.2     27      >50   ND    >50  ND   ND                              49      1.0     >50     >50   ND    >50  ND   ND                              50      2.5     >50     >50   ND    >50  ND   ND                              51      0.9     50      33    ND    >50  ND   ND                              52      4.3     22      34    ND    ND   ND   ND                              53      0.4     12      50    ND    >50  ND   ND                              54      1.1     25      >50   ND    >50  ND   ND                              55      2.5     5.3     >50   ND    >50  ND   ND                              56      0.7     6.5     >50   ND    ND   ND   ND                              57      0.5     25      >50   ND    ND   ND   ND                              58      0.2     6       50    ND    ND   ND   ND                              59      0.3     12      25    ND    ND   ND   ND                              60      >50     >50     >50   ND    ND   ND   ND                              61      25      1.9     0.5   ND    ND   ND   ND                              62      50      50      >50   ND    ND   ND   ND                              63      <50     >50     40    ND    ND   ND   ND                              64      >50     >50     >50   ND    ND   ND   ND                              65      >50     >50     >50   ND    ND   ND   ND                              ______________________________________                                         ND = Not determined.                                                     

Cell Culture

PDGF Receptor Autophosphorylation Assay

Rat aorta smooth muscle cells (RASMC) are isolated from the thoracicaorta of rats and explanted according to the method of Ross, J. Cell.Biol., 30: 172-186 (1971). Cells are grown in Dulbecco's modifiedEagle's medium (DMEM, Gibco) containing 10% fetal calf serum (FBS,Hyclone, Logan, Utah), 1% glutamine (Gibco) and 1%penicillin/streptomycin (Gibco). Cells are identified as smooth musclecells by their "hill and valley" growth pattern and by fluorescentstaining with a monoclonal antibody specific for SMC μ-actin (Sigma).RASMC are used between passages 5 and 20 for all experiments. Testcompounds are prepared in dimethylsulfoxide (DMSO) in order to achieveconsistency in the vehicle and to ensure compound solubility.Appropriate DMSO controls are simultaneously evaluated with the testcompounds.

Rat aortic smooth muscle cells are grown to confluencey in 100 mmdishes. Growth medium is removed and replaced with serum-free medium andcells are incubated at 37° C. for an additional 24 hours. Test compoundsare then added directly to the medium and cells incubated for anadditional 2 hours. After 2 hours PDGF-BB is added at a finalconcentration of 30 ng/mL for 5 minutes at 37° C. to stimulateautophosphorylation of PDGF receptors. Following growth factortreatment, the medium is removed, and cells are washed with coldphosphate-buffered saline and immediately lysed with 1 mL of lysisbuffer (50 mM Hepes, pH 7.5, 150 mM NaCl, 10% glycerol, 1% Triton-X 100,1 mM EDTA, 1 mM EGTA, 50 mM NaF, 1 mM sodium orthovanadate, 30 mMp-nitrophenyl phosphate, 10 mM sodium pyrophosphate, 1 mM phenylmethylsulfonyl fluoride, 10 μg/mL aprotinin and 10 μg/mL leupeptin). Lysatesare centrifuged at 10,000 ×g for 10 minutes. Supernatants are incubatedwith 10 μL of rabbit anti human PDGF type AB receptor antibody (1:1000)for 2 hours. Following the incubation, protein-A-sepharose beads areadded for 2 hours with continuous mixing, and immune complexes bound tothe beads washed 4 times with 1 mL lysis wash buffer. Immune complexesare solubilized in 40 μL of Laemmli sample buffer and electrophoresed in8-16% SDS polyacrylamide gels. Following electrophoresis, separatedproteins are transferred to nitrocellulose and immunoblotted with a1:1000 dilution of antiphosphotyrosine monoclonal antibody (UBI clone4G10; #05-321). Following extensive washing with PBS-0.2% tween-20, theblots are incubated with horseradish peroxidase labeled goat antimouseIgG (1:5000; Bio-Rad Inc, Hercules, Calif.) and protein levels aredetected by enhanced chemiluminescence (ECL) detection system accordingto the instructions of the supplier (Amersham Inc, Arlington Heights,Ill.). The density of the protein bands are determined using NIH Imagesoftware (v. 1.56) and IC₅₀ values are generated from the densitometricdata.

Table 2 provides data for inhibition of PDGF-stimulated receptorautophospharylation in rat aorta smooth muscle cells.

                  TABLE 2                                                         ______________________________________                                        Example Number  IC.sub.50 (μM)                                             ______________________________________                                        8               >10                                                           11              >10                                                           13              2                                                             38              0.7                                                           40              >10                                                           45              3.5                                                           59              2.2                                                           ______________________________________                                    

Human Colon Carcinoma Growth Delay Assay

Human colon carcinoma cells are seeded into 96-well tissue cultureplates in a final volume of 180 μL of 10% fetal bovine serum containinggrowth media and allowed to incubate overnight (37° C., 5% CO₂, 95%air). Cells of the SW620 cell line are seeded at 1.0-1.5×10⁴ cells perwell. Cells of the HCT8 and HT29 cell lines are seeded at 2-4×10³ cellsper well. Serially diluted drug solutions are prepared in growth mediumat 10 ×concentration; 20 μL of these solutions are added to duplicatewells and incubated with the cells for 3 days in a cell cultureincubator. At the end of the incubation period, cells are fixed with 100μL per well of 10% trichloroacetic acid after removing the drug/culturemedium. The plates are washed 5 times with tap water and stained with100 μL per well of 0.075% sulforhodamine B in 1% acetic acid for 10minutes. The plates are rinsed 4 times and allowed to air dry. The stainin the wells are solubilized by the addition of 10 mM unbuffered Trisbase and the absorbance read using a microtiter plate optical reader.Inhibition of cell growth is calculated from absorbance data of thetreated cells compared to untreated control cells.

Human Colon Carcinoma Clonogenic Assay

Human colon carcinoma cells are seeded into 6 well plates in volumes of3 mL and allowed to incubate overnight (37° C., 5% CO₂, 95% air). SW620cells are seeded at 7×10⁵ per well; HCT8 cells are seeded at 5×10⁵ perwell; HT29 cells are seeded at 4×10⁵ cells per well. Serially diluteddrugs are prepared at 200 ×the final concentration and 15 μL are addedto each of duplicate wells. Cells are incubated with drug for 2 days,rinsed once with 1 mL of trypsin+EDTA, and then trypsinized with thesame trypsin solution. After trituration and centrifugation at 750 ×gfor 3 minutes, the cells are suspended in serum-free growth medium andcounted using an electronic particle counter. An agarose mixtureappropriate for the cloning of each cell line is made using 10% fetalbovine serum in growth medium (SW620-0.35% agarose, HCT8 and HT29-0.4%agarose). An appropriate volume of medium containing the drug treatedcells is suspended into the agarose-serum mixture to give final cellconcentrations in 2.5 mL of 1.75×10⁴ SW620, 1.25×10⁴ HCT8, and 7.5×10³HT29. One milliliter of each of these cell suspensions is added toduplicate wells of 6 well plates previously prepared with 10%serum/growth medium/ 1% agarose plugs. The cells in these plates areincubated for approximately 2 weeks in the incubator and stained with 1mL per well of 1 mg/mL iodonitrotetrazolium violet stain. The visiblecolonies are counted with an electronic optical colony counter and theclonogenicity of treated cells calculated in comparison to untreatedcontrol cells.

Table 3 provides data for inhibition of growth in human adenocarcinomacell lines.

                                      TABLE 3                                     __________________________________________________________________________    Cell Line (IC.sub.50, μM)                                                  Example                                                                            HCT-8  SW-620 HT-29  HCT-8 SW-620                                                                              HT-29                                   Number                                                                             Growth Delay                                                                         Growth Delay                                                                         Growth Delay                                                                         Clonogenic                                                                          Clonogenic                                                                          Clonogenic                              __________________________________________________________________________    11   ND     >25    ND     ND     2.1, >5                                                                            ND                                      13   ND     >25    ND     ND         >5                                                                             ND                                      59   ND     >25    ND     ND        >25                                                                             ND                                      __________________________________________________________________________     ND = Not determined.                                                     

What is claimed is:
 1. A method of treating protein tyrosine kinase mediated cancer, the method comprising administering to a patient having protein tyrosine kinase mediated cancer a therapeutically effective amount of a compound of Formula I ##STR16## wherein Ar is aryl, or substituted aryl;R¹, R², R³, and R⁴ are each independently hydrogen, C₁ -C₆ alkyl, --OC₁ -C₆ alkyl, --OH, halogen, --CO₂ R⁴, --CONR^(a) R^(b), --NO₂, --NR^(a) R^(b), --COC₁ -C₆ alkyl, --CHO, --CN, --SO₂ C₁ -C₆ alkyl, ##STR17## --OCH₂ --CH(OH)CH₂ OH, --O(CH₂)_(n) NR^(a) R^(b), --SH, --SC₁ -C₆ alkyl, or --S(CH₂)_(n) --NR^(a) R^(b), or R² and R³ together can form a cycloalkyl ring and R^(a) and R^(b) are each independently hydrogen or C₁ -C₆ alkyl, n is 0 to 5, or a pharmaceutically acceptable salt, ester, amide, or prodrug thereof.
 2. A method of treating or preventing restenosis, the method comprising administering to a patient having restenosis or at risk of having restenosis a therapeutically effective amount of a compound of Formula I ##STR18## wherein Ar is aryl, or substituted phenyl wherein the phenyl substituent is halogen, C₁ -C₈ alkyl, --CN, --CF₃, --NO₂, --NH₂ --NHC₁ -C₈ alkyl, --N(C₁ -C₈ alkyl)₇, --OC₁ -C₈ alkyl, or --OHR¹, R², R³, and R⁴ are each independently hydrogen, C₁ -C₆ alkyl, --OC₁ -C₆ alkyl, --OH, halogen, --CO₂ R², --CONR^(a) R^(b), --NO₂, --NR^(a) R^(b), --COC₁ -C₆ alkyl, --CHO, --CN, --SO₂ C₁ -C₆ alkyl, ##STR19## --OCH₂ CH(OH)CH₂ OH, --O(CH₂)_(n) NR^(a) R^(b), --SH, --SC₁ -C₆ alkyl, or --S(CH₂)_(n) --NR^(a) R^(b), or R² and R³ together can form a cycloalkyl ring; and R^(a) and R^(b) are each independently hydrogen or C₁ -C₆ alkyl, n is 0 to 5, or a pharmaceutically acceptable salt, ester, amide, or prodrug thereof.
 3. A method of treating atherosclerosis, the method comprising administering to a patient having atherosclerosis a therapeutically effective amount of a compound of Formula I ##STR20## wherein Ar is aryl, or substituted phenyl wherein the phenyl substituent is halogen, C₁ -C₈ alkyl, --CN, --CF₃, --NO₂, --NH₂ --NHC₁ -C₈ alkyl, --N(C₁ -C₈ alkyl)₂, --OC₁ -C₈ alkyl, or --OHR¹, R², R³, and R⁴ are each independently hydrogen, C₁ -C₆ alkyl, --OC₁ -C₆ alkyl, --OH, halogen, --CO₂ R^(a), --CONR^(a) R^(b), --NO₂, --NR^(a) R^(b), --COC₁ -C₆ alkyl, --CHO, --CN, --SO₂ C₁ -C₆ alkyl, ##STR21## --OCH₂ CH(OH)CH₂ OH, --O(CH₂)_(n) NR^(a) R^(b), --SH, --SC₁ -C₆ alkyl, or --S(CH₂)_(n) --NR^(a) R^(b), or R² and R³ together can form a cycloalkyl ring; and R^(a) and R^(b) are each independently hydrogen or C₁ -C₆ alkyl, n is 0 to 5, or a pharmaceutically acceptable salt, ester, amide, or prodrug thereof.
 4. A method of treating psoriasis, the method comprising administering to a patient having psoriasis a therapeutically effective amount of a compound of Formula I ##STR22## wherein Ar is aryl, or substituted aryl;R¹, R², R³, and R⁴ are each independently hydrogen, C₁ -C₆ alkyl, --OC₁ -C₆ alkyl, --OH, halogen, --CO₂ R², --CONR^(a) R^(b), --NO₂, --NR^(a) R^(b), --COC₁ -C₆ alkyl, --CHO, --CN, --SO₂ C₁ -C₆ alkyl, ##STR23## --OCH₂ CH(OH)CH₂ OH, --O(CH₂)_(n) NR^(a) R^(b), --SH, --SC₁ -C₆ alkyl, or --S(CH₂)_(n) --NR^(a) R^(b), or R² and R³ together can form a cycloalkyl ring; and R^(a) and R^(b) are each independently hydrogen or C₁ -C₆ alkyl, n is 0 to 5, or a pharmaceutically acceptable salt, ester, amide, or prodrug thereof.
 5. A method of inhibiting protein tyrosine kinases, the method comprising administering to a patient in need of protein tyrosine kinases inhibition a protein tyrosine kinase inhibiting amount of a compound of Formula I ##STR24## wherein Ar is aryl, or substitued aryl, heteroaryl;R¹, R², R³, and R⁴ are each independently hydrogen, C₁ -C₆ alkyl, --OC₁ -C₆ alkyl, --OH, halogen, --CO₂ R^(a), --CONR^(a) R^(b) 2, --NO₂, --NR^(a) R^(b), --COC₁ -C₆ alkyl, --CHO, --CN, --SO₂ C₁ -C₆ alkyl, ##STR25## --OCH₂ CH(OH)CH₂ OH, --O(CH₂)_(n) NR^(a) R^(b), --SH, --SC₁ -C₆ alkyl, or --S(CH₂)_(n) --NR^(a) R^(b), or R² and R³ together can form a cycloalkyl ring; and R^(a) and R^(b) are each independently hydrogen or C₁ -C₆ alkyl, n is 0 to 5, or a pharmaceutically acceptable salt, ester, amide, or prodrug thereof.
 6. The method of claim 5 wherein the protein tyrosine kinase is FGFr.
 7. The method of claim 5 wherein the protein tyrosine kinase is PDGFr.
 8. A method of treating or preventing restenosis, the method comprising administering to a patient having restenosis or at risk of having restenosis a therapeutically effective amount of the compound 1-(3-formlphenyl)benzimidazole or a pharmaceutically acceptable salt, ester, amide, or prodrug thereof.
 9. A method of treating atherosclerosis, the method comprising administering to a patient having atherosclerosis a therapeutically effective amount of the compound 1-(3-formlphenyl)benzimidazole or a pharmaceutically acceptable salt, ester, amide or prodrug thereof. 